PROGNOSTIC VALUE OF

A Dissertation on

PROGNOSTIC VALUE OF

HYPOCHOLESTEROLEMIA IN SEPSIS

Submitted to

THE TAMILNADU DR. M.G.R. MEDICAL UNIVERSITY CHENNAI – 600032

In partial fulfilment of the Regulations for the Award of the Degree of

M.D. BRANCH - I GENERAL MEDICINE

DEPARTMENT OF GENERAL MEDICINE STANLEY MEDICAL COLLEGE

CHENNAI – 600 001

APRIL - 2017

CERTIFICATE BY THE INSTITUTION

This is to certify that Dr.K.RAJ SANTAN, Post - Graduate Student (May 2014 TO April 2017) in the Department of General Medicine STANLEY MEDICAL COLLEGE, Chennai- 600 001, has done this dissertation on

“Prognostic value of Hypocholesterolemia in sepsis” under my guidance and supervision in partial fulfillment of the regulations laid down by the Tamilnadu Dr. M. G. R. Medical University, Chennai, for M.D. (General Medicine), Degree Examination to be held in April 2016.

Dr.P.Vasanthi, M.D. Dr.Dr.Isaac Christian Moses, M.D , FICP , FACP.,

Professor and HOD Dean

Department of Medicine, Govt. Stanley Medical College Govt. Stanley Medical College & Hospital.

& Hospital.

CERTIFICATE BY THE GUIDE

This is to certify that Dr.K.RAJ SANTAN, Post - Graduate Student (MAY 2014 TO APRIL 2017) in the Department of General Medicine STANLEY MEDICAL COLLEGE, Chennai- 600 001, has done this dissertation on “Prognostic value of Hypocholesterolemia in sepsis” under my guidance and supervision in partial fulfillment of the regulations laid down by the Tamilnadu Dr. M. G. R. Medical University, Chennai, for M.D. (General Medicine), Degree Examination to be held in April 2016.

DR. S. ASHOK KUMAR, M.D.

Associate Professor, Department of Medicine,

Govt. Stanley Medical College & Hospital, Chennai – 600001.

DECLARATION

IDr.K.RAJSANTAN,, declare that I carried out this work on

“Prognostic value of Hypocholesterolemia in sepsis” at the intensive medical care unit of Government Stanley Hospital . I also declare that this bonafide work or a part of this work was not submitted by me or any other for any award, degree, or diploma to any other university, board either in India or abroad.

This is submitted to The Tamilnadu DR. M. G. R. Medical University, Chennai in partial fulfilment of the rules and regulation for the M. D. Degree examination in General Medicine.

Dr.K.RAJ SANTAN

ACKNOWLEDGEMENT

At the outset I thank our dean Dr. ISAAC CHRISTIAN MOSES, M.D, FICP, FACP for permitting me to carry out this study in our hospital.

I express my profound thanks to my esteemed Professor and Teacher Dr. P. VASANTHI, M.D., Professor and HOD of Medicine, Stanley Medical College Hospital, for encouraging and extending invaluable guidance to perform and complete this dissertation.

I immensely thank my unit chief Dr. ASHOK KUMAR, M.D., D.T.C.D, Associate Professor Of Medicine for his constant encouragement and guidance throughout the study. I would also like to extend my heartfelt thanks to my former unit chief Dr. G. VASUMATHI M.D., for her guidance in completing this dissertation.

I wish to thank Dr. A.RAMALINGAM M.D, and Dr. S. PRAKASH, M.D, Assistant Professors of my unit, Department of Medicine, Stanley Medical College Hospital for their valuable suggestions, encouragement and advice.

I sincerely thank the members of Institutional Ethical Committee, Stanley Medical College for approving my dissertation topic.

I thank all my colleagues, House Surgeons, and Staff nurses and other para medical workers for their support.

At this juncture I would also want to extend my heartfelt gratitude to my parents and my wife for the motivation and encouragement extended by them which gave fulfilment to the dissertation work.

I sincerely thank all those Patients who participated in this study, for their co-operation.

Above all, I thank the Almighty for gracing me this opportunity, health, and knowledge throughout this study.

PROGNOSTIC VALUE OF

HYPOCHOLESTEROLEMIA IN

SEPSIS

CONTENTS

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BIBILIOGRAPHY

PROFORMA

CONSENT FORM

ETHICAL COMMITTEE APPROVAL LETTER

MASTER CHART

ABBREVIATIONS

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INTRODUCTION

“Sepsis is the most common cause of death in Intensive Care Unit. It is an increasingly common cause of mortality and morbidity in elderly, Immunocompromised and critically ill patients. Approximately, 25–35% of patients with severe sepsis and 40–55% of patients with septic shock die within 30 days.

Patients with systemic infection and organ dysfunction are often difficult to distinguish from patients with similar clinical signs and lab findings, but without infection. The established biological markers of inflammation (leukocytes, C-reactive protein) may be influenced by parameters other than infection and may only be slowly released during progression of an infection . Despite aggressive management, sepsis continues to have a high mortality rate as high as 48.8% . The need for an early prognostic marker to identify those at highest risk for mortality in order to optimize treatment is essential”

“Procalcitonin (PCT) and C-reactive protein (CRP) are the present standard markers in the ICU setting; however, these test carry significant limitations. Cholesterol may be a useful prognostic marker in sepsis because lipid metabolism is significantly altered by systemic inflammatory response.

These changes occur within hours of an inflammatory response and are negatively correlated to clinical outcome. Lipopolysaccharide (LPS), the major component of the outer membrane of Gram-negative bacteria, plays a key role

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in the initiation of sepsis. After LPS binds to CD14, the inflammatory cascade begins. Also, there is a significant transition in the distribution of circulating lipoproteins before and after sepsis. Previous studies illustrated that lipoprotein plays an important role in LPS binding and neutralization, enzyme incorporation, including paraoxonase and platelet-activating factor acetylhydrolase, inhibition of the expression of endothelial cell adhesion, and stimulation of the expression of endothelial nitric oxide synthase in vitro . Patients with severe sepsis have low levels of cholesterol, including high- density lipoprotein (HDL), low-density lipoprotein (LDL), and apolipoprotein A-I (Apo A-I) and high levels of triglycerides (TGs) and free fatty acids (FFAs)”

The objective of this study is to asses whether hypocholesterolemia can be used as a prognostic indicator in sepsis

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ubiquinone and oxysterols are synthesizedvia certain intermediate steps in cholesterolbiosynthesis.

Cholesterol is an important component in thecell membrane. It is essential for the appropriatemembrane composition needed by a certain celltype. Signal transduction is especially intense inthe areas where cholesterol and sphingolipids formspecial areas of membrane communication, namelymembrane rafts. This membrane cholesterol is inequilibrium with the unesterified cholesterol in the

plasma pool. Cholesterol is carried mainly by thelow-density lipoprotein (LDL) fraction in the plasma

EXOGENOUS PATHWAY OF LIPID METABOLISM:

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“Lipoprotein metabolism can be divided into exogenous and endogenous pathways. The exogenous pathway starts with the absorption of dietary cholesterol and fatty acids from intestine. The mechanisms regulating the amount of dietary cholesterol that is absorbed are unknown. Sitosterolemia is a rare autosomal recessive disorder associated with hyperabsorption of cholesterol and plant sterols from the intestine . These genes involved are expressed in the liver and intestine and are upregulated by cholesterol feeding;

they may normally cooperate to limit intestinal sterol absorption .

Within the intestinal cell, free fatty acids combine with glycerol to form triglycerides, and cholesterol is esterified by acyl-coenzyme A:cholesterol acyltransferase (ACAT) to form cholesterol esters. The important role of ACAT was established in an animal model of ACAT deficiency, which found complete resistance to diet-induced hypercholesterolemia due to lack of cholesterol ester synthesis and reduced capacity to absorb cholesterol . Despite this, clinical trials have found that ACAT inhibitors may worsen atherosclerosis . Triglycerides and cholesterol are assembled intracellularly as chylomicrons. The main apolipoprotein is B-48, but C-II and E are acquired as the chylomicrons enter the circulation. Apo B-48 permits lipid binding to the chylomicron but does not bind to the low density lipoprotein receptor, thereby preventing premature clearance of chylomicrons from the circulation before they are acted upon by lipoprotein lipase (LPL)”

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Apo C-II is a cofactor for LPL that makes the chylomicrons progressively smaller, primarily by hydrolyzing the core triglycerides and releases free fatty acids. The free fatty acids are then used as source of energy, converted to triglyceride, or stored in adipose tissue. The end-products of chylomicron metabolism are chylomicron remnants which are cleared from the circulation by hepatic chylomicron remnant receptors for which apo E is a high-affinity ligand. The chylomicron remnants contain a smaller core of lipids which are enveloped by excess surface components. These surface constituents are transferred from the chylomicron remnant and forms high density lipoprotein.

ENDOGENOUS PATHWAY OF LIPID METABOLISM :

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“ The endogenous pathway of lipid metabolism begins with the synthesis of very low density lipoprotein (VLDL) by the liver . VLDL particles contain a core of triglycerides (60 percent by mass) and cholesterol esters (20 percent by mass). Microsomal triglyceride transfer protein (MTP) is an intracellular lipid-transfer protein found in the endoplasmic reticulum. It is essential for the transfer of the lipid molecules (principally triglycerides) onto apolipoprotein (apo) B 100 in the liver . The surface apolipoproteins for VLDL are noted above. They include apo C-II, which acts as a cofactor for lipoprotein lipase; apo C-III, which inhibits this enzyme; and apo B-100 and E, which serve as ligands for the apolipoprotein B/E (low density lipoprotein [LDL]) receptor . In the absence of functional MTP, VLDL is not secreted into the circulation. Abetalipoproteinemia is a rare genetic disorder in which MTP is absent. “

“The triglyceride core of nascent VLDL particles is hydrolyzed by lipoprotein lipase. During lipolysis, the core of the VLDL particle is reduced, generating VLDL remnant particles (also called intermediate density lipoprotein [IDL]) that are depleted of triglycerides via a process similar to the generation of chylomicron remnants. Some of the excess surface components in the remnant particle, including phospholipid, unesterified cholesterol, and apolipoproteins A, C and E, are transferred to high density lipoprotein (HDL).

VLDL remnants can either be cleared from the circulation by the apo B/E (LDL) or the remnant receptors, or remodeled by hepatic lipase to

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form LDL particles. There are four common sequence polymorphisms in the hepatic lipase gene promoter; the most frequent is a C to T substitution . The presence of a C allele is associated with higher hepatic lipase activity; smaller, denser, and more atherogenic LDL particles; and inversely with lower levels of HDL cholesterol .”

Low density lipoprotein:

“ LDL particles contain a core of cholesterol esters, lesser amounts of triglyceride, and are enriched in apolipoprotein B-100, which is the ligand for binding to the apo B/E (LDL) receptor. LDL can be internalized by hepatic and nonhepatic tissues. Hepatic LDL cholesterol can be converted to bile acids and secreted into the intestinal lumen. LDL cholesterol is internalized by nonhepatic tissues and can be used for hormone production, cell membrane synthesis, or stored in esterified form.

The internalization of LDL is regulated by cellular cholesterol requirements via negative feedback control of apo B/E (LDL) receptor expression . Cells in positive cholesterol balance, for example, suppress apo B/E (LDL) receptor expression. On the other hand, decreased activity of HMG CoA reductase, the enzyme that controls the rate of de novo cholesterol synthesis by the cell, leads ultimately to a fall in cell cholesterol, increased expression of apo B/E (LDL) receptors, enhanced uptake of cholesterol from the circulation, and a decrease in the plasma cholesterol concentration.”

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Chemically-modified LDL, such as oxidized LDL, can enter macrophages and some other tissues through unregulated scavenger receptor.

This pathway results in excess accumulation of intracellular cholesterol and forms foam cells, which leads to the formation of atheromatous plaques.

The importance of the LDL receptor in the regulation of cholesterol metabolism has been demonstrated in both experimental animals and humans.

Knockout of the LDL receptor in transgenic mice lead to a significant elevation in total cholesterol levels, a defect that can be reversed by restoring the LDL receptor gene . In humans, familial hypercholesterolemia is associated with a defect in the LDL receptor gene.

High density lipoprotein:

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“ The formation and metabolism of HDL particles involves the following steps :

●Hepatic and intestinal synthesis of small nascent HDL particles composed of phospholipid and apolipoproteins.

●Procurement of surface components (phospholipids, cholesterol, and apolipoproteins) from triglyceride-depleted chylomicron and VLDL remnants.

●Acquisition of free (unesterified) cholesterol from tissue sites (such as the liver and macrophages in the arterial wall) and other lipoproteins; the initial HDL particles contain relatively little cholesterol.

HDL particles are thought to participate in cholesterol metabolism in the following way:

●Nascent HDL particles (cholesterol-absent and phospholipid-depleted) promote the transfer of intracellular cholesterol to the cell membrane , in the peripheral tissues, through the action of protein known as ABCA1 . ABCA1 expression on the cell surface is induced by cholesterol loading and reduced after the cholesterol is removed by apolipoproteins. Mutations in the gene encoding for ABCA1 is associated with low serum HDL-cholesterol concentrations in familial HDL deficiency and Tangier disease. Apo A-I on the surface of HDL plays a key role in this process by signaling a transduction protein to mobilize cholesterol esters from intracellular pools.

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One common variant of this gene, R219K, is associated with higher HDL-C and lower triglyceride concentrations; carriers of this variant have a reduced severity of coronary disease, a slower progression of disease, and fewer coronary events . Other genetic variations in the ABCA1 gene may also contribute to determining HDL-C concentrations in the population.

●After acquisition of free cholesterol by the HDL particle, the cholesterol is esterified to cholesterol esters by lecithin:cholesterol acyl transferase (LCAT), a plasma enzyme that is activated primarily by apo A-I.

By a similar mechanism, HDL can act as an acceptor for cholesterol which is released during lipolysis of triglyceride-containing lipoproteins.

●Lipid transfer proteins, such as cholesteryl ester transfer protein facilitates movement of the cholesterol esters to apo B-containing lipoproteins (VLDL, IDL, and LDL). This cholesterol can then be delivered to the tissues for steroid synthesis or storage or to the liver for the conversion into bile acids”

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Cholesteryl ester transfer protein:

• The role of cholesteryl ester transfer protein (CETP) in lipoprotein metabolism is complex, and the impact of CETP on cardiovascular disease is not well understood

• Circulating CETP mediates the transfer of cholesteryl esters from HDL particles to the triglyceride-rich lipoproteins LDL and VLDL; at the same time, triglycerides are transferred in the opposite direction.

• In this process, HDL-cholesterol is decreased, the cholesterol content in VLDL is increased, and LDL particles become smaller and denser.

• Intracellular CETP in both the periphery and the liver appears to promote cholesterol removal from peripheral cells and uptake by the liver

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Lipoprotein(a):

• Lipoprotein(a) or Lp(a) is a specialized form of LDL that is assembled extracellularly from apolipoprotein (a) and LDL . Apo(a) links to apolipoprotein B-100 on the surface of LDL by disulfide bridges.

• The formation of apo(a):apo B complexes requires an LDL particle of a certain morphology and composition. The structural integrity of LDL, and therefore Lp(a) formation, are modulated by LCAT .

• The apo(a) chain contains five domains known as kringles . The fourth kringle contains regions that are homologous with the fibrin-binding domains of plasminogen. Through this structural similarity to plasminogen, Lp(a) interferes with fibrinolysis by competing with plasminogen binding to plasminogen receptors, fibrinogen, and fibrin.

• The net effect is impairment in plasminogen activation and plasmin generation at the thrombus surface, leading to decreased thrombolysis .Lp(a) can also bind to macrophages via a high-affinity receptor, possibly promoting foam cell formation and localization of Lp(a) in atherosclerotic plaques .

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SEPSIS:

“ Sepsis is a clinical syndrome that has physiologic, biologic, and biochemical abnormalities caused by a dysregulated inflammatory response to infection. Sepsis and the inflammatory response that ensues can lead to multiple organ dysfunction syndrome and death

The incidence of sepsis varies among the different racial and ethnic groups, but appears to be highest among African-American males .

The incidence is also greatest during the winter, probably due to the increased prevalence of respiratory infections .

Older patients ≥65 years of age account for the majority (60 to 85 percent) of all episodes of sepsis; with an increasing aging population, it is likely that the incidence of sepsis will continue to increase in the future”

DEFINITION:

A 2016 SCCM/EISCM task force has defined sepsis as “life-threatening organ dysfunction caused by a dysregulated host response to infection”

Organ dysfunction :

“Organ dysfunction is defined by the 2016 SCCM/ESICM task force as an increase of two or more points in the SOFA score. The validity of this score was derived from critically-ill patients with suspected sepsis by interrogating

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over a million intensive care unit (ICU) electronic health record encounters from ICUs both inside and outside the United States . ICU patients were suspected as having infection if body fluids were cultured and they received antibiotics. Predictive scores (SOFA, systemic inflammatory response syndrome [SIRS], and logistic Organ Dysfunction System [LODS]) were compared for their ability to predict mortality. Among critically ill patients with suspected sepsis, the predictive validity of the SOFA score for in-hospital mortality was superior to that for the SIRS criteria (area under the receiver operating characteristic curve 0.74 versus 0.64). Patients who fulfill these criteria have a predicted mortality of ≥10 percent. Although the predictive capacity of SOFA and LODS were similar, SOFA is considered easier to calculate, and was therefore recommended by the task force”

PATHOPHYSIOLOGY OF SEPSIS:

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The normal host response to an infection is a complex process that localises and controls bacterial invasion, with repair of injured tissue. It includes the activation of circulating and fixed phagocytic cells, as well as the generation of proinflammatory and antiinflammatory mediators. Sepsis results when the response to infection becomes generalized and involves normal tissues which are remote from the site of injury or infection.

NORMAL RESPONSE TO INFECTION:

“The host response to an infection is initiated when innate immune cells, particularly macrophages, recognize and bind to microbial antigens. This may occur by many pathways:

●Pattern recognition receptors (PRRs) on the surface of host immune cells binds to the pathogen-associated molecular patterns (PAMPs) of microorganisms . There are three families of PRRs: toll-like receptors (TLRs), nucleotide-oligomerization domain (NOD) leucine-rich repeat proteins, and retinoic-acid-inducible gene I (RIG-I)-like helicases. Examples include the peptidoglycan of Gram-positive bacteria binding to TLR-2 on host immune cells, as well as the lipopolysaccharide of Gram-negative bacteria binding to TLR-4 and/or lipopolysaccharide-binding protein (CD14 complex) on host immune cells.

●PRRs can also recognize endogenous danger signals, so-called alarmins or danger-associated molecular patterns (DAMPs) which are released

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during the inflammatory response. DAMPs are nuclear, cytoplasmic, or mitochondria structures acquiring new functions when released in to the extracellular environment. Examples of DAMPs include high mobility group box-1 protein HMGB1, S100 proteins, and mitochondrial DNA .

●The triggering receptor expressed on myeloid cell (TREM-1) and the myeloid DAP12-associating lectin (MDL-1) receptors on host immune cells may recognize and bind to microbial components .

The binding of immune cell surface receptors to microbial components have multiple effects:

●The engagement of TLRs elicits a signaling cascade via the activation of cytosolic nuclear factor-kb (NF-kb). Activated NF-kb moves from the cytoplasm to the nucleus, binds to transcription sites, and induces activation of a large set of genes involved in the host inflammatory response, such as proinflammatory cytokines (tumor necrosis factor alpha [TNFa], interleukin-1 [IL-1]), chemokines (intercellular adhesion molecule-1 [ICAM-1], vascular cell adhesion molecule-1 [VCAM-1]), and nitric oxide.

●Polymorphonuclear leukocytes (PMNs) becomes activated and express adhesion molecules that cause their aggregation and margination in the vascular endothelium. This is facilitated by the endothelium expressing adherence molecules to attract leukocytes. The PMNs then go through a series of steps (rolling, adhesion, diapedesis, and chemotaxis) to migrate to the site of

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injury . The release of mediators by PMNs at the site of infection is responsible for the cardinal signs of local inflammation: warmth and erythema due to local vasodilation and hyperemia, and protein-rich edema due to increased microvascular permeability.

This process is highly regulated by a mixture of proinflammatory and antiinflammatory mediators secreted by macrophages, which have been triggered and activated by the invasion of tissue by bacteria :

●Proinflammatory mediators – Important proinflammatory cytokines include TNFa and interleukin-1 (IL-1), which share a remarkable array of biological effects . The release of TNFa is self-sustaining (ie, autocrine secretion), while non-TNF cytokines and mediators (eg, Il-1, IL-2, IL-6, IL-8, IL-10, platelet activating factor, interferon, and eicosanoids) increase the levels of other mediators (ie, paracrine secretion). The proinflammatory milieu leads to the recruitment of more PMNs and macrophages.

●Antiinflammatory mediators – Cytokines which inhibit the production of TNFa and IL-1 are considered antiinflammatory cytokines. Such antiinflammatory mediators suppress the immune system by inhibiting the cytokine production by mononuclear cells and monocyte-dependent T helper cells. However, their effects may not be universally antiinflammatory. As for example, IL-10 and IL-6 both enhance B cell function (proliferation, immunoglobulin secretion) and encourage the development of cytotoxic T cells .

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The balance of proinflammatory and antiinflammatory mediators regulates the inflammatory processes, including adherence, chemotaxis, phagocytosis of the invading bacteria, bacterial killing, and phagocytosis of debris from injured tissue. If the mediators balance each other and the initial infectious insult is overcome, homeostasis will be restored . The end result will be tissue repair and healing”

TRANSITION TO SEPSIS:

“Sepsis occurs when the release of proinflammatory mediators in response to an infection exceeds the boundaries of the local environment, leading to a more generalized response . When a similar process occurs in response to a noninfectious condition (eg, pancreatitis, trauma), the process is referred to as systemic inflammatory response syndrome (SIRS).

Sepsis can be conceptualized as malignant intravascular inflammation ,

1. Malignant because it is uncontrolled, unregulated, and self-sustaining 2. Intravascular because the blood spreads mediators that are usually

confined to cell-to-cell interactions within the interstitial space

3. Inflammatory because all characteristics of the septic response are exaggerations of the normal inflammatory response

It is uncertain why immune responses that usually remains localized sometimes spread beyond the local environment causing sepsis. The cause is

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likely multifactorial and may include the direct effects of the invading microorganisms or their toxic products, release of large quantities of proinflammatory mediators, and complement activation. In addition, some individuals may be genetically susceptible to developing sepsis”

Effects of microorganism:

Bacterial cell wall components (endotoxin, peptidoglycan,lipoteichoic acid and muramyl dipeptide) and bacterial products (staphylococcal enterotoxin B, Pseudomonas exotoxin A ,toxic shock syndrome toxin-1, and M protein of hemolytic group A streptococci) contribute to the progression of a local infection to sepsis . This is supported by the following observations regarding endotoxin, a lipopolysaccharide found in the cell wall of gram negative bacteria:

●Endotoxin is detectable in the blood of septic patients.

●Elevated plasma levels of endotoxin are associated with shock and multiple organ dysfunction.

●Endotoxin reproduces many of the features of sepsis when it is infused into humans, including activation of the complement, coagulation, and fibrinolytic systems . These effects may lead to microvascular thrombosis and the production of vasoactive products, such as bradykinin.

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Excess proinflammatory mediators:

“ Large quantities of proinflammatory cytokines released in patients with sepsis may spill into the bloodstream, contributing to the progression of a local infection to sepsis. These include tumor necrosis factor alpha (TNFa) and interleukin-1 (IL-1), whose plasma levels peak early and eventually decrease to undetectable levels. Both cytokines can cause fever, hypotension, leukocytosis, induces other proinflammatory cytokines, and the simultaneous activation of coagulation and fibrinolysis .

The evidence indicating that TNFa has an important role in sepsis is particularly strong. It includes the following:

• Circulating levels of TNFa are higher in septic patients than non-septic patients with shock

• Infusion of TNFa produces symptoms similar to those observed in septic shock

• Anti-TNFa antibodies protect animals from lethal challenge with endotoxin

. The high levels of TNFa in sepsis are due in part to the binding of endotoxin to lipopolysaccharide (LPS)-binding protein and its subsequent transfer to CD14 on macrophages, which stimulates TNFarelease ”

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Complement activation:

“The complement system is a protein cascade that helps clear pathogens from an organism .There is evidence that activation of the complement system plays an important role in sepsis; most importantly, inhibition of the complement cascade decreases inflammation and improves mortality in animal models:

● In a rodent model of sepsis, a complement fragment 5a receptor (C5aR) antagonist decreased mortality, inflammation, and vascular permeability . In contrast, increased production of complement fragment 5a (C5a) and increased expression of C5aR enhanced neutrophil trafficking .

● In several animal models of sepsis (lipopolysaccharide injection in mice and rats, Escherichia coli infusion in dogs and baboons, and cecal

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ligation and puncture in mice), a complement fragment 1 (C1) inhibitor decreased mortality, inflammation, and vascular permeability”

Genetic susceptibility:

The single nucleotide polymorphism (SNP) is the most common form of genetic variation. SNPs are stable substitutions of a single base that have a frequency of more than one percent in at least one population and are strewn throughout the genome, including promoters and intergenic regions. At most, only 2 to 3 percent alter the function or expression of a gene. The total number of common SNPs in the human genome is estimated to be more than 10 million. SNPs are used as genetic markers.

Various SNPs are associated with increased susceptibility to infection and poor outcomes. They include SNPs of genes that encode cytokines (eg, TNF, lymphotoxin-alpha, IL-10, IL-18, IL-1 receptor antagonist, IL-6, and interferon gamma), cell surface receptors (eg, CD14, MD2, toll-like receptors 2 and 4, and Fc-gamma receptors II and III), lipopolysaccharide ligands (lipopolysaccharide binding protein, bactericidal permeability increasing protein), mannose-binding lectin, heat shock protein 70, angiotensin I- converting enzyme, plasminogen activator inhibitor, and caspase-12 .

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SYSTEMIC EFFECTS OF SEPSIS:

Widespread cellular injury occurs when the immune response becomes generalized; cellular injury is the precursor to organ dysfunction. The precise mechanism of cellular injury is not understood, but its occurrence is indisputable as autopsy studies have shown widespread endothelial and parenchymal cell injury.

Mechanisms proposed to explain the cellular injury include:

• Tissue ischemia (insufficient oxygen relative to oxygen need)

• Cytopathic injury (direct cell injury by proinflammatory mediators and/or other products of inflammation)

• Altered rate of apoptosis (programmed cell death).

Tissue ischemia:

“Significant derangement in metabolic autoregulation, the process that matches oxygen availability to changing tissue oxygen needs, is typical of sepsis.

In addition, microcirculatory and endothelial lesions frequently develop during sepsis. These lesions reduce the cross-sectional area available for tissue oxygen exchange, disrupting tissue oxygenation and causing tissue ischemia and cellular injury:

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●Microcirculatory lesions – The microcirculatory lesions may be the result of imbalances in the coagulation and fibrinolytic systems, both of which are activated during sepsis.

●Endothelial lesions – The endothelial lesions may be a consequence of interactions between endothelial cells and activated polymorphonuclear leukocytes (PMNs). The increase in receptor-mediated neutrophil-endothelial cell adherence induces the secretion of reactive oxygen species, lytic enzymes, and vasoactive substances (nitric oxide, endothelin, platelet-derived growth factor, and platelet activating factor) into the extracellular milieu, whichinjures the endothelial cells. Lipopolysaccharide (LPS) may also induce cytoskeleton disruption and microvascular endothelial barrier integrity, in part, through nitric oxide synthase (NOS), Ras homolog gene family member A (RhoA), and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) activation .

Another factor contributing to tissue ischemia in sepsis is that erythrocytes lose their normal ability to deform within the systemic microcirculation . Rigid erythrocytes have difficulty navigating the microcirculation during sepsis, causing excessive heterogeneity in the microcirculatory blood flow and depressed tissue oxygen flux”

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Cytopathic injury :

“ Proinflammatory mediators and/or other products of inflammation may cause sepsis-induced mitochondrial dysfunction (eg, impaired mitochondrial electron transport) via a variety of mechanisms, including direct inhibition of respiratory enzyme complexes, oxidative stress damage, and mitochondrial DNA breakdown . Such mitochondrial injury leads to cytotoxicity. There are several lines of evidence that support this belief:

●Cell culture experiments have shown that endotoxin, tumor necrosis factor alpha (TNFa), and nitric oxide cause destruction and/or dysfunction of inner membrane and matrix mitochondrial proteins, followed by degeneration of the mitochondrial ultrastructure. These changes are followed by measurable changes in other cellular organelles by several hours . The end result is functional impairment of mitochondrial electron transport, disordered energy metabolism, and cytotoxicity.

●Studies using various animal models have found normal or supranormal oxygen tension in organs during sepsis, suggesting impaired oxygen utilization at the mitochondrial level. As examples, a study in resuscitated endotoxemic pigs found a supranormalileomucosal oxygen tension , while a study in endotoxemic rats found an elevated oxygen tension in the bladder epithelium .

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The clinical relevance of mitochondrial dysfunction in septic shock was suggested by a study of 28 critically ill septic patients who underwent skeletal muscle biopsy within 24 hours of admission to the intensive care unit (ICU) . Skeletal muscle adenosine triphosphate (ATP) concentrations, a marker of mitochondrial oxidative phosphorylation, were significantly lower in the 12 patients who died of sepsis than in 16 survivors. In addition, there was an association between nitric oxide overproduction, antioxidant depletion, and severity of clinical outcome. Thus, cell injury and death in sepsis may be explained by cytopathic (or histotoxic) anoxia, which is an inability to utilize oxygen even when present.

Mitochondria can be repaired or regenerated by a process called biogenesis. Mitochondrial biogenesis may prove to be an important therapeutic target, potentially accelerating organ dysfunction and recovery from sepsis”

Apoptosis:

“Apoptosis (also called programmed cell death) describes a set of regulated physiologic and morphologic cellular changes leading to cell death.

This is the principal mechanism by which senescent cells are normally eliminated and the dominant process by which inflammation is terminated once an infection has subsided.

During sepsis, proinflammatory cytokines may delay apoptosis in activated macrophages and neutrophils, thereby prolonging or augmenting the

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inflammatory response and contributes to the development of multiple organ failure. Sepsis also induces extensive lymphocyte and dendritic cell apoptosis, which alters the immune response efficacy and results in decreased clearance of invading microorganisms. Apoptosis of lymphocytes has been observed at autopsies in both animal and human sepsis.

The extent of lymphocyte apoptosis correlates with the severity of the septic syndrome and the level of immunosuppression. Apoptosis has been also observed in parenchymal cells, endothelial, and epithelial cells. Several animal experiments show that inhibiting apoptosis protects against organ dysfunction and lethality”.

Mitochondrial dysfunction in sepsis-induced multiple organ failure:

“ In patients dying from sepsis, light and electron microscopy as well as immunohistochemical staining for markers of cellular injury and stress, revealed that cell death was rare in sepsis-induced cardiac and renal dysfunction.

Moreover, the degree of cell injury or death did not account for severity of sepsis-induced organ dysfunction . The presence of subtle mitochondrial morphological changes could indicate that mitochondrial energetic crisis (metabolic substrate utilization and mitochondrial OxPhos machinery perturbations) may be involved in organ dysfunction, in the absence of cell death”.

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Immunosuppression:

“Clinical observations and animal studies suggest that the excess inflammation of sepsis may be followed by immunosuppression . Among the evidence supporting this hypothesis, an observational study removed the spleens and lungs from 40 patients who died with active severe sepsis and then compared them with the spleens from 29 control patients and the lungs from 30 control patients . The median duration of sepsis was four days. The secretion of proinflammatory cytokines (ie, tumor necrosis factor, interferon gamma, interleukin-6, and interleukin-10) from the splenocytes of patients with severe sepsis was generally less than 10 percent that of controls, following stimulation with either anti-CD3/anti-CD28 or lipopolysaccharide. Moreover, the cells from the lungs and spleens of patients with severe sepsis exhibited increased expression of inhibitory receptors and ligands, as well as expansion of suppressor cell populations, compared with cells from control patients. The inability to secrete proinflammatory cytokines combined with enhanced expression of inhibitory receptors and ligands suggests clinically relevant immunosuppression.”

ORGAN-SPECIFIC EFFECTS OF SEPSIS:

The cellular injury described above, accompanied by the release of proinflammatory and antiinflammatory mediators, often progresses to organ dysfunction. No organ system is protected from the consequences of sepsis.

Multiple organ dysfunction is common.

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Circulation:

“Hypotension due to diffuse vasodilation is the most severe expression of circulatory dysfunction in sepsis. It is probably an unintended consequence of the release of vasoactive mediators, whose purpose is to improve metabolic autoregulation (the process that matches oxygen availability to changing tissue oxygen needs) by inducing appropriate vasodilation. Mediators include the vasodilators prostacyclin and nitric oxide (NO), which are produced by endothelial cells.

NO is believed to play a central role in the vasodilation accompanying septic shock, since NO synthase can be induced by incubating vascular endothelium and smooth muscle with endotoxin . When NO reaches the systemic circulation, it depresses metabolic autoregulation at all of the central, regional, and microregional levels of the circulation. In addition, NO may trigger an injury in the central nervous system that is localized to areas that regulate autonomic control .

Another factor that may contribute to the persistence of vasodilation during sepsis is impaired compensatory secretion of antidiuretic hormone (vasopressin). This hypothesis is supported by a study that found that plasma vasopressin levels were lower in patients with septic shock than in patients with cardiogenic shock (3.1 versus 22.7 pg/mL), even though the groups had similar systemic blood pressures . It is also supported by numerous small studies that

31

demonstrated that vasopressin improves hemodynamics and allows other pressors to be withdrawn .

Vasodilation is not the only cause of hypotension during sepsis.

Hypotension may also be due to redistribution of intravascular fluid. This is a consequence of both increased endothelial permeability and reduced arterial vascular tone leading to increased capillary pressure.

In addition to these diffuse effects of sepsis on the circulation, there are also localized effects:

●In the central circulation (ie, heart and large vessels), decreased systolic and diastolic ventricular performance due to the release of myocardial depressant substances is an early manifestation of sepsis . Despite this, ventricular function may still be able to use the Frank Starling mechanism to increase cardiac output, which is necessary to maintain the blood pressure in the presence of systemic vasodilation.

Patients with preexisting cardiac disease (eg, elderly patients) are often unable to increase their cardiac output appropriately.

●In the regional circulation (ie, small vessels leading to and within the organs), vascular hyporesponsiveness (ie, inability to appropriately vasoconstrict) leads to an inability to appropriately distribute systemic blood flow among organ systems. As an example, sepsis interferes with the

32

redistribution of blood flow from the splanchnic organs to the core organs (heart and brain) when oxygen delivery is depressed [57].

●The microcirculation (ie, capillaries) may be the most important target in sepsis. Sepsis is associated with a decrease in the number of functional capillaries, which causes an inability to extract oxygen maximally.

Techniques including reflectance spectrophotometry and orthogonal polarization spectral imaging have allowed in vivo visualization of the sublingual and gastric microvasculature . Compared to normal controls or critically ill patients without sepsis, patients with severe sepsis have decreased capillary density . This may be due to extrinsic compression of the capillaries by tissue edema, endothelial swelling,and/or plugging of the capillary lumen by leukocytes or red blood cells (which lose their normal deformability properties in sepsis).

●At the level of the endothelium, sepsis induces phenotypic changes to endothelial cells. This occurs through direct and indirect interactions between the endothelial cells and components of the bacterial wall. These phenotypic changes cause endothelial dysfunction, which is associated with coagulation abnormalities, reduced leukocytes, impaired red blood cell deformability, upregulation of adhesion molecules, adherence of platelets and leukocytes, and degradation of the glycocalyxstructure . Diffuse endothelial activation leads to widespread tissue edema, which is rich in protein.

33

Microparticles from circulating and vascular cells also participate in the deleterious effects of sepsis-induced intravascular inflammation”.

Lung :

“Endothelial injury in the pulmonary vasculature during sepsis disturbs the capillary blood flow and enhances microvascular permeability, resulting in interstitial and alveolar pulmonary edema. Neutrophil entrapment within the lung's microcirculation initiates the injury in the alveolocapillary membrane.

The result is pulmonary edema, which creates ventilation-perfusion mismatch and leads to hypoxemia. Such lung injury is prominent during sepsis, likely reflecting the lung's large microvascular surface area. Acute respiratory distress syndrome is a manifestation of these effects”.

Gastrointestinal tract:

The circulatory abnormalities typical of sepsis may depress the gut's normal barrier function, allowing translocation of bacteria and endotoxin into the systemic circulation (possibly via lymphatics, rather than the portal vein) and extending the septic response . This is supported by animal models of sepsis, as well as a prospective cohort study that found that increased intestinal permeability (determined from the urinary excretion of orally administered lactuloseand mannose) was predictive of the development of multiple organ dysfunction syndrome .

34

Liver:

“The reticuloendothelial system of the liver normally acts as the first line of defense in clearing bacteria and bacteria-derived products that have entered the portal system from the gut. Liver dysfunction can prevent the elimination of enteric-derived endotoxin and bacteria-derived products, which precludes the appropriate local cytokine response and permits direct spillover of these potentially injurious products into the systemic circulation”.

Kidney :

“ Sepsis is often accompanied by acute renal failure. The mechanisms by which sepsis and endotoxemia lead to acute renal failure are incompletely understood. Acute tubular necrosis due to hypoperfusion and/or hypoxemia is one mechanism . However, systemic hypotension, direct renal vasoconstriction, release of cytokines (eg, tumor necrosis factor [TNF]), and activation of neutrophils by endotoxin and FMLP (a three amino acid [fMet-Leu-Phe]

chemotactic peptide in bacterial cell walls) may also contribute to renal injury”.

“Growing evidence suggest that septic acute renal failure is only in part sustained by renal hypoperfusion. It has been shown that sepsis is associated with normal or even elevated renal blood flow, which is associated with redistribution of blood flow from cortical to medullary region. These macrovascular changes are associated with microcirculatory dysfunction, inflammatory response induced by pathogen-associated molecular patterns

35

(PAMPs) and danger-associated molecular patterns (DAMPs) and bio- energetic adaptation response including tubular cell cycle arrest machinery.

Hence, the mechanism of kidney injury during sepsis may be viewed as a bio- energetics adaptation of tubular epithelial cells induced by deregulated inflammation in response to peritubular microvascular dysfunction.

The role of renal replacement therapy (RRT) in septic patients has been evaluated both for renal support and immunomodulation . Retrospective clinical studies have suggested that early initiation of RRT and use of continuous methods are associated with a better hemodynamic tolerance and outcome . Timing and dose of RRT are ongoing sources of debate, yet available randomized clinical trials fails to demonstrate any beneficial impact .likelihood of death is increased in patients with sepsis who develop renal failure, It is not well understood why this occurs”.

Nervous system :

“Central nervous system (CNS) complications occur frequently in septic patients, often before the failure of other organs. The most common CNS complications are an altered sensorium (encephalopathy). The pathogenesis of the encephalopathy is poorly defined. A high incidence of brain microabscesses was noted in one study, but the significance of hematogenous infection as the principal mechanism remains uncertain because of the heterogeneity of the observed pathology.

36

CNS dysfunction has been attributed to changes in metabolism and alterations in cell signalling due to inflammatory mediators. Dysfunction of the blood brain barrier probably contributes, allowing increased leukocyte infiltration, exposure to toxic mediators, and active transport of cytokines across the barrier. Mitochondrial dysfunction and microvascular failure both precede functional CNS changes, as measured through somatosensory evoked potentials .

In addition to these neurological consequences of sepsis, there is growing recognition that the parasympathetic nervous system may be a mediator of systemic inflammation during sepsis. This is supported by numerous observations in various animal models. Afferent vagus nerve stimulation during sepsis increases the secretion of corticotropin-releasing hormone (CRH), ACTH, and cortisol; the last effect can be suppressed by subdiaphragmaticvagotomy. Parasympathetic tone affects thermoregulation, as experimental vagotomy attenuates thehyperthermic response to IL-1 . Efferent parasympathetic activity, mediated by acetylcholine, has an antiinflammatory effect on the cytokine profile, with decreased in vitro expression of the proinflammatory cytokines TNF, interleukin (IL)-1, IL-6 and IL-18 . And, external vagal stimulation prevents the onset of shock following vagotomy, while an acetylcholine receptor agonist diminishes the pathologic response to sepsis”

37

SOFA SCORE:

“The SOFA score was initially designed to sequentially assess the severity of organ dysfunction in patients who were critically ill from sepsis,Since multi organ dysfunction is common in critically ill patients, it has since been used to predict mortality in those with organ failure from other causes including those with acute liver failure from acetaminophenoverdose, chronic liver failure (CLIF-SOFA), and cancer, as well as in patients who have undergone cardiac surgery or hematopoietic stem cell transplant .

SOFA uses simple measurements of major organ function to calculate a severity score, The scores are calculated 24 hours after admission to the ICU and every 48 hours thereafter (thus, the term "Sequential" Organ Failure Assessment). The mean and the highest scores are most predictive of mortality.

In addition, scores that increase by about 30 percent are associated with a mortality of at least 50 %.

The SOFA severity score is based upon the following measurements of organ function:

1. Respiratory system – the ratio of arterial oxygen tension to fraction of inspired oxygen (PaO2/FiO2)

2. Cardiovascular system – the amount of vasoactive medication necessary to prevent hypotension

3. Hepatic system – the bilirubin level

38

4. Coagulation system – the platelet concentration 5. Neurologic system – the Glasgow coma score 6. Renal system – the serum creatinine or urine output

The SOFA score has been endorsed by the Society of Critical Care Medicine (SCCM) and the European Society of Intensive Care Medicine (ESICM) as a tool to facilitate the identification of patients at risk of dying from sepsis”

●Sepsis :

“ Sepsis is now defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. As an organ dysfunction score, SOFA can be used to identify those whose organ dysfunction is "life- threatening" such that an increase in the SOFA score ≥2 is associated with a mortality of ≥10 percent”

●Septic shock :

“ Patients with a SOFA score ≥2 who also have a vasopressor requirement and an elevated lactate >2 mmol/L (>18 mg/dL) despite adequate fluid resuscitation have a predicted mortality of 40 percent.

The validity of this score was derived from millions of ICU electronic health record encounters both inside and outside the United States. Among critically ill patients with suspected sepsis, the predictive validity of the SOFA

39

score for in-hospital mortality was superior to that for the systemic inflammatory response criteria

Importantly, despite the SCCM/ESICM endorsement of the SOFA score, many experts caution clinicians regarding the use of SOFA. The SOFA score does not diagnose sepsis, identify those whose organ dysfunction is truly due to infection, or determine individual treatment strategies or individual outcome. Rather, the SOFA score helps identify patients, as a group, who potentially have a high risk of death from infection”

QUICK SOFA:

• “The quick SOFA (qSOFA) score has also been proposed by the SCCM/ESICM as a tool to help identify patients with early sepsis outside of the ICU.

• Patients are assigned one point each for the following clinical features which can be easily measured at the bedside: respiratory rate ≥22/min, altered mentation, and systolic blood pressure ≤100 mmHg.

• Patients with two or more of these features were reported to have a poor outcomes from sepsis.

• The qSOFA requires additional prospective validation before it can be routinely used to identify those at risk of death from sepsis outside the ICU”

SO

HYPOCH

“Ac cholestero hypochole

FA SCOR

HOLESTE

ccording t l levels b esterolemia

RE:

EROLEMI

to the A below 160 a”

40

IA:

American H mg/dL or

Heart Ass r 4.1 mmo

sociation ol/l are to

in 1994, be classi

total ified as

41

CAUSES OF DECREASED LEVELS OF CHOLESTEROL ARE:

• Malnutrition

• Malabsorption

• Liver disease

• Hyperthyroidism

• Anaemia

• Sepsis

• Stress

• statin

• Antibiotics

• Adrenal insufficiency

• Pernicious anaemia

• Haemolytic jaundice

• Hyperthyroidism

• Severe infections

• leukemia

• Terminal stages of malignancy

• Hypolipoproteinemias

• Manganese deficiency

42

HYPOCHOLESTEROLEMIA IN SEPSIS:

It is well established that serum lipid concentrations are decreased during sepsis.The mechanism of hypocholesterolemia during infection is multifactorial, due to decreased synthesis and increased catabolism playing a role. Decreased lipoprotein synthesis occurs in vitro when hepatocytes are exposed to TNF and IL-6. Infection and inflammation induce oxidation of LDL-cholesterol. Decreases in HDL-cholesterol may be related to high concentrations of phospholipase A2, or due to downregulation of the ATP- bindingcassette transporter-1 gene.Hypolipidemia reduces competition for binding of LPS to lipopolysaccharide-binding protein (LBP), leading to ligation of the CD14 complex and activation of mononuclear cells. Conversely, binding of LPS to lipoproteins facilitates delivery of LPS to hepatocytes for detoxification, which if insufficient may lead to increased mononuclear cell activation.Several studies highlight the relationship between low serum cholesterol and sepsis

43

AIMS AND OBJECTIVES OF STUDY

The aim of this study was to examine the difference and dynamic changes in serum lipid levels in ICU patients with sepsis and evaluate whether these lipids are associated with prognosis

44

MATERIALS AND METHODS

PLACE OF STUDY :

Intensive Medical Care Unit

Department of General Medicine.

Govt.Stanley Medical College & hospital

STUDY DESIGN: Cross sectional observational study

STUDY POPULATION: 100

STUDY PERIOD: March 2016 to August 2016

INCLUSION CRITERIA:

1. Age ≥ 18 years

2. patients admitted to the ICU with sepsis between March and August 2016

45

EXCLUSION CRITERIA:

1. age less than 18 years,

2. pregnancy,

3. ICU readmission, and a

4. need for primary cardiac care.

5. Patients with liver disease (hepatitis B, hepatitis C, immune hepatitis, liver cirrhosis, and hepatocellular carcinoma),

6. Dyslipidemia

7. history of statin or steroid (≥15 mg/day) use within the previous 7 days

METHODOLOGY:

For each patient, data on age, gender, comorbidity, administration of vasoactive drugs, length of ICU stay, and 28-day mortality were collected.

In addition, Sequential Organ Failure Assessment (SOFA) scores, and laboratory data, including cholesterol, TG, HDL, LDL were collected on days 0 (ICU admission)and day 3.

46

A detailed case report form/ proforma is to be used for data collection of the patient which includes the details mentioned.

OPERATIONAL DEFINITION:

“Sepsis was clinically defined as the identification of an infection site and three or four systemic inflammatory response syndrome (SIRS) criteria: (1) body temperature less than 36°C (96.8°F) or greater than 38°C (100.4°F); (2) heart rate greater than 90beats/min; (3) tachypnea of greater than 20breaths/min or an arterial partial pressure of carbon dioxide less than 4.3kPa (32mmHg); and (4) white blood cell count of less than 4000cells/mm³ (4 × 10⁹cells/L) or greater than 12,000cells/mm³ (12 × 10⁹cells/L)”

Septic shock was defined as the state of sepsis with persistent hypotension despite adequate volume resuscitation

HUMAN SUBJECT PROTECTION:

• The full protocol along with draft questionnaire and Informed consent will be kept in Institutional ethical Committee and approval will be obtained.

47

INFORMED CONSENT:

• Consent form will be written in both English and Tamil and consent will be obtained from the participant, confidentiality will be maintained.

48

RESULTS AND DISCUSSIONS

Groups Definition Number

Group A • Total

Cholesterol < 160 mg/dl

53

Group B • Total

Cholesterol ≥ 160 mg/dl

47

49

Null Hypothesis

Null Hypothesis : H0 Total Cholesterol < 160 mg/dl group equal in effect compared to Total Cholesterol ≥ 160 mg/dl group

Alternate Hypothesis : H1 Total Cholesterol < 160 mg/dl group hazardous in effect compared to Total Cholesterol ≥ 160 mg/dl group

Data Analysis

Descriptive statistics was done for all data and were reported in terms of mean values and percentages. Suitable statistical tests of comparison were done. Continuous variables were analysed with the unpaired t test.. Categorical variables were analysed with the Chi-Square Test and Fisher Exact Test.

Statistical significance was taken as P < 0.05. The data was analysed using SPSS version 16 and Microsoft Excel 2007.

51

Age Distribution Group A Group B Combined

Mean 55.47 52.79 54.21

SD 9.16 11.26 10.24

P value

Unpaired t Test

0.1922

Among the study patients, there was no statistically significant difference in relation to age distribution between group A (mean=55.47, SD=9.16) and group B (mean=52.79, SD=11.26) with a p value of <0.05 as per unpaired t test. Therefore we fail to reject the null hypothesis that there is no difference in age distribution between the study groups.

Ag

≤ 30 y

31-50 51-70

> 70 y Total

ge Gr

years 1 years 10 years 42 years 0

53

1 2 3 4 5 6 7 8

roup A

1

0 1

2 2

2

3 4

1 0

0 0 0 0 0 0 0 0

Group B

1 19 25 2 47

10 42

Group A

≤ 30 year

50

Combin

2 29 67 2 100

0 1

rs 31-50 y

ned Gro ( 1.89 18.87 79.25 0.00 100

19 25

Group B

Age

years 51-7

oup A (%)

2

7 4

5 5

4 1

2

70 years >

Group B (%) 2.13 40.43 53.19 4.26 100

2 29

Combin

> 70 years

Combin (%) 2.00 29.00 67.00 2.00 100

67

2 ned

ned

Gender

Male Female Total P value Fishers Ex

0 10 20 30 40 50 60

Group A

22 31 53

xact Test

22

Group

A Group

26 21 47

31

A

52

p B Com

48 52 100

26

Gr

Gen

Male

mbined G

4 58 10 0

21

roup B

der

Female

Group A (%) 1.51 8.49 00 .1677

4

Group B (%) 55.32 44.68 100

48

52

Combined

B Comb (%

48.00 52.00 100

bined

%)

Am difference 58.49%) a as per unp no differen

0.0 20.0 40.0 60.0 80.0 100.0 120.0 140.0

Mean Values

mong the in relation and group B

aired t test nce in gend

102.94 11702

00 00 00 00 00 00 00 00

SBP (m

study pa n to gender B (majorit t. Therefore der status b

6842

117.02 110.90

mm Hg) D

G

53

atients, the r status bet ty are male e we fail to between the

68.42 74.47 71.63

BP (mm Hg)

Vital Pa

Group A G

ere was tween grou es – 55.32%

o reject the e study gro

103.06 94.64

PR (bpm

aramete

Group B C

no statisti up A (majo

%)) with a e null hypot

oups.

27.23

98.08

m) RR (br m

ers

ombined

ically sign ority are fem a p value o

thesis that

23.66 24.97

reaths per min)

Te

nificant males – of <0.05 there is

99.61 99.68 99.62

emperature (F)

54

Vital Parameters SBP

(mm Hg)

DBP (mm Hg)

PR (bpm)

RR (breaths per min)

Temperature (F)

Group A Mean 102.94 68.42 103.06 27.23 99.61

SD 26.53 14.31 18.26 10.55 1.14

Group B Mean 117.02 74.47 94.64 23.66 99.68

SD 22.45 12.30 12.31 8.16 0.73

Combined Mean 110.90 71.63 98.08 24.97 99.62

SD 25.21 13.35 15.74 9.32 0.93

P value

Unpaired t Test

0.0054 0.0264 0.0089 0.0446 0.7276

SBP

Among the study patients, there was a statistically significant difference in relation to systolic blood pressure distribution between group A (mean=102.94, SD=26.53) and group B (mean=117.02, SD=22.45) with a p value of <0.05 as per unpaired t test. Therefore we reject the null hypothesis that there is no difference in systolic blood pressure distribution between the study groups.

55

Discussion

The mean SBP was significantly less in group A compared to group B by a mean difference of 14.08mm Hg (12% lower). This difference is significant with a p-value of 0.0054 as per unpaired t test.

DBP

Among the study patients, there was a statistically significant difference in relation to diastolic blood pressure distribution between group A (mean=68.41, SD=14.31) and group B (mean=74.47, SD=12.30) with a p value of <0.05 as per unpaired t test. Therefore we reject the null hypothesis that there is no difference in diastolic blood pressure distribution between the study groups.

Discussion

The mean DBP was significantly less in group A compared to group B by a mean difference of 6.05mm Hg (8% lower). This difference is significant with a p-value of 0.0264 as per unpaired t test.

56

PR

Among the study patients, there was a statistically significant difference in relation to respiratory rate distribution between group A (mean=103.06, SD=18.26) and group B (mean=94.64, SD=12.31) with a p value of <0.05 as per unpaired t test. Therefore we reject the null hypothesis that there is no difference in respiratory rate distribution between the study groups.

Discussion

The mean respiratory rate was significantly more in group A compared to group B by a mean difference of 8.42 bpm (8% higher). This difference is significant with a p-value of 0.0089 as per unpaired t test.

RR

Among the study patients, there was a statistically significant difference in relation to respiratory rate distribution between group A (mean=27.23, SD=10.55) and group B (mean=23.66, SD=8.16) with a p value of <0.05 as per unpaired t test. Therefore we reject the null hypothesis that there is no difference in respiratory rate distribution between the study groups.

57

`Discussion

The mean respiratory rate was significantly more in group A compared to group B by a mean difference of 3.57 breaths per min (13% higher). This difference is significant with a p-value of 0.0446 as per unpaired t test.

Temperature

Among the study patients, there was a statistically significant difference in relation to temperature distribution between group A (mean=99.61, SD=1.14) and group B (mean=99.68, SD=0.73) with a p value of <0.05 as per unpaired t test. Therefore we reject the null hypothesis that there is no difference in temperature distribution between the study groups.

0.00 10.00 20.00 30.00 40.00 50.00 60.00 70.00

Mean Values 5860

0 0 0 0 0 0 0 0

58.60 41.49

Urea (mg

Re

Gro

58

48.85

g/dl)

enal Par

up A Grou

rameter

up B Com

1.64

Creatinin

rs

bined

1.17 1.37

ne (mg/dl)