Biomedical subjects

I H Chaudry

Publications and source records attributed to I H Chaudry.

At least 109 records · Page 6Linked to original sources

PubMed1999

Organ dysfunction following hemorrhage and sepsis: mechanisms and therapeutic approaches (Review).

Despite significant advances in the management of trauma victims, sepsis and the ensuing multiple organ failure remain the leading causes of death in the surgical intensive care unit. Although much effort has been focused on the mediators released in large quantities following shock and sepsis, blockade of mediators such as proinflammatory cytokines has not yet resulted in a successful therapy. However, as more studies are forthcoming, the mechanisms responsible for cell and organ dysfunctions following trauma-hemorrhage and sepsis are becoming better understood, and promising new therapeutic approaches are currently being evaluated. In order to understand the precise mechanisms responsible for cellular dysfunction and consequently irreversible organ damage and multiple organ failure, it is important to correlate various pathophysiological changes with mediators and signal transduction pathways at the cellular and subcellular level. In this review we focus first on factors and mediators responsible for producing cell and organ dysfunctions, especially hepatocellular dysfunction, following trauma, hemorrhagic shock, and sepsis. The changes in signaling transduction pathways will also be discussed, specifically the role of mitogen-activated protein kinases, transcription factors, nitric oxide, heat shock proteins, and inflammatory cytokines in the development of cell and organ dysfunctions following trauma-hemorrhage and sepsis. Moreover, potential therapeutic approaches for improving cell and organ functions under adverse circulatory conditions are included.

Adenosine Triphosphate↗

PubMed1999

Immunological consequences of trauma and shock.

The immune system is a powerful, complex entity composed of numerous cell types and regulated by autocrine, paracrine, and hormonal mechanisms. Trauma and haemorrhagic shock induce numerous changes within this system which are ultimately deleterious and contribute to the high incidence of organ dysfunction and infectious complications seen following injury. Regional hypoxia and depletion of intracellular energy stores occur in response to diminished microcirculatory blood flow, and these changes alter cellular signalling and result in the release of pro-inflammatory cytokines and prostanoids which mediate further suppression of immune function. Neutrophil priming serves to induce tissue damage in critical organ systems such as the lungs, heart, liver, and gut, further insulting the injured organism. Depression of antigen presentation and cytokine elaboration by macrophages and other antigen presenting cells effectively prevents a normal response from the acquired immune system, and lymphocyte-monocyte interactions are squelched. The resulting depression in cell mediated and humoral immunity renders the organism susceptible to microbial infection and contributes to the morbidity and mortality associated with nosocomial infections. Hormonal modulation of the immune response is highly evident following trauma and haemorrhage, and the preponderance of male morbidity associated with sepsis can be explained by the depression in immune function seen in males, but not females in the pro-oestrous state. Despite the multitude of changes induced by trauma and haemorrhage, experimental studies have revealed several promising pharmacologic interventions which may serve to blunt the effect of injury on the immune system, and render the host competent to withstand the bacterial and viral challenges responsible for so much of the late mortality following severe injury.

Adjuvants, Immunologic↗

PubMed1999

Trauma-hemorrhage delays wound healing potentially by increasing pro-inflammatory cytokines at the wound site.

BACKGROUND: Studies indicate impaired wound healing after trauma. The underlying mechanism remains unknown. METHODS: Mice were subjected to midline laparotomy, and polyvinyl alcohol sponges were implanted subcutaneously before hemorrhage (35 +/- 5 mmHg for 90 minutes, resuscitated) or sham operation. Wound exudate cells from the sponges were harvested on the first, third, and fifth postoperative day and cultured for 24 hours. Interleukin (IL)-1 beta, IL-6, monocyte chemotactic protein-1 (MCP-1), macrophage inflammatory protein-2 (MIP-2), and transforming growth factor (TGF)-beta were determined in the supernatants. IL-1 beta and IL-6 were measured in the wound fluid. RESULTS: Hemorrhage decreased collagen deposition in the wound. TGF-beta release was significantly decreased on the first and third postoperative days after hemorrhage, whereas IL-1 beta and IL-6 release was increased at 3 and 5 days after hemorrhage. Similarly, IL-1 beta and IL-6 in the wound fluid were significantly increased at 3 days after hemorrhage. CONCLUSIONS: Because increased levels of pro-inflammatory cytokines and decreased amounts of TGF-beta have been reported to impair the process of wound healing, the increased release of IL-1 beta and IL-6 and the decreased release of TGF-beta after hemorrhage might contribute to the decreased collagen production in those animals. Thus, attempts to locally change the ratio of those cytokines in trauma victims might be useful for improving wound healing in those patients.

Animals↗

PubMed1999

What is the role of interleukin 10 in polymicrobial sepsis: anti-inflammatory agent or immunosuppressant?

BACKGROUND: Controversy exists concerning the role of interleukin 10 (IL-10) in sepsis. When IL-10 is used in models of endotoxemia, it appears to protect (by anti-inflammatory effects), whereas in models of polymicrobial sepsis it seems to be deleterious (by immunosuppression?). However, little direct evidence for such an immunosuppressive role is available for polymicrobial sepsis. Thus the aim of this study was to determine whether IL-10 contributes to lymphocyte immunosuppression in a model of cecal ligation and puncture (CLP) and whether neutralization of IL-10 has any salutary effects on survival after sepsis. METHODS: To assess the former, polymicrobial sepsis was induced in male C57BL/6J wild-type (+/+) and C57BL/6J-IL-10 knockout(-/-) mice by CLP. Splenocytes were harvested 24 hours later and stimulated with concanavalin A to assess their proliferative capacity and their ability to release the Th1 lymphokines interleukin 2 and interferon gamma (by enzyme-linked immunosorbent assay, nanograms/millilter). To further verify the immunosuppressive role of IL-10, splenocytes were obtained from male C3H/HeN mice 24 hours after CLP and then stimulated in the presence or absence of anti-IL-10 monoclonal antibody (Mab, 4 micrograms/mL). To assess the in vivo effects of IL-10 neutralization on survival after CLP, C3H/HeN mice (16 per group) were given 250 micrograms of anti-IL-10 Mab (intraperitoneally) either immediately after CLP (before the initiation of the hyperdynamic phase) or 12 hours after CLP (the beginning of the hypodynamic state). Control mice were given nonspecific rat immunoglobulin G. RESULTS: These data indicate that IL-10 deficiency (-/-) prevents the depression of the proliferative capacity and Th1 lymphokine production after sepsis. Analysis of the interleukin 2-interferon gamma production patterns and proliferative capacity in lymphocytes treated with anti-IL-10 Mab confirmed the role of IL-10 in suppressing lymphocyte responsiveness in CLP. Interestingly, however, only delayed administration (12 hours after CLP) of anti-IL-10 markedly increased survival of mice (Fisher's exact test, P < .05). CONCLUSION: The results not only illustrate IL-10's role in septic immune dysfunction but document that anti-IL-10 administration beyond the initial proinflammatory hyperdynamic state of polymicrobial sepsis improves survival of animals subjected to sepsis.

Animals↗

PubMed1998

Upregulation of Kupffer cell beta-adrenoceptors and cAMP levels during the late stage of sepsis.

Although a burst of immunoresponsiveness may occur during the early stage of sepsis, late sepsis is characterized by severe immunodepression. In addition, although studies have shown that stimulation of macrophage beta-adrenoceptors results in an increase in cAMP and an associated reduction in macrophage phagocytic activity, it remains unknown whether Kupffer cell beta-adrenoceptor characteristics and cAMP levels are altered during polymicrobial sepsis. To study this, Sprague-Dawley rats were subjected to sepsis by cecal ligation and puncture (CLP). At 5 h (i.e., the early stage of sepsis) or 20 h (late sepsis) after CLP or sham operation, the liver was perfused with collagenase solution and Kupffer cells were isolated. beta-Adrenoceptor characteristics of the isolated Kupffer cells were determined using [125I]iodopindolol, and basal levels of cAMP were measured by radioimmunoassay. The results indicate that while maximum binding capacity (Bmax) of Kupffer cell beta-adrenoceptors was not altered at 5 h, it increased significantly at 20 h after CLP. Similarly, basal levels of cAMP in Kupffer cells did not change at 5 h but increased markedly at 20 h after the onset of sepsis. In contrast, the dissociation constant (Kd, 1/affinity) of Kupffer cell beta-adrenoceptors was not significantly affected by sepsis at both 5 h and 20 h after CLP. Thus, upregulation of beta-adrenoceptors and increase in cAMP levels in Kupffer cells occur during the late stage of polymicrobial sepsis, and this may contribute to the depression of macrophage phagocytic function under such conditions.

Adrenergic beta-Antagonists↗

PubMed1998

Increased mucosal B-lymphocyte apoptosis during polymicrobial sepsis is a Fas ligand but not an endotoxin-mediated process.

Sepsis is reported to induce an increase in the rate of apoptosis (Ao), in immature lymphoid cells residing in hematopoietic tissues such as the thymus and bone marrow. Alternatively, secondary lymphoid tissue, such as the spleen exhibit little innate (unstimulated) Ao. However, it is unknown whether or not polymicrobial sepsis has any effects on the frequency of Ao in mucosal lymphoid tissue and what, if any, are the functional consequences of such a change. To assess this, Peyer's patch cells were harvested from C3H/HeN (endotoxin-sensitive) mice killed 12 or 24 hours after the onset of polymicrobial sepsis (cecal ligation and puncture [CLP]). The results indicate that the percentage of cells that were Ao+ as determined by flow cytometry were markedly increased at 24 hours, but not at 12 hours post-CLP. This correlates well with evidence of increased DNA fragmentation as well as histological changes observed both at a light and transmission electron microscopic level of the Peyer's patch Ao. Phenotypically, these changes were restricted to the B220+ (B-cell) population that also exhibited a marked increase of Fas/Apo-1 antigen expression. The functional consequence of this increased apoptosis appears to be associated with the endogenous stimulation (activation) of IgA production by mucosal B lymphocytes and increased nuclear c-Rel expression. Furthermore, we found that Peyer's patch lymphocytes isolated from C3H/HeJ-Faslgld (endotoxin-tolerant/Fas ligand- [FasL] deficient) as opposed to C3H/HeJ (endotoxin-tolerant) inbred mice did not exhibit increased Ao after CLP. These findings indicate that increased B-cell Ao appears to be a FasL-Fas antigen-mediated process, but is not due to endotoxin sensitivity. In conclusion, we speculate that the increased Fas-associated apoptosis detected in mucosal B cells (as opposed to splenic or bone marrow B cells) may be due to increased luminal antigens other than endotoxin, released due to gut barrier integrity breakdown during sepsis.

Animals↗

PubMed1998

Is gut the "motor" for producing hepatocellular dysfunction after trauma and hemorrhagic shock?

BACKGROUND: Although studies suggest that the gut may be the "motor" responsible for producing sepsis and multiple organ failure after injury, it is not known whether enterectomy prior to the onset of hemorrhage alters proinflammatory cytokines TNF and IL-6 and, if so, whether hepatocellular dysfunction and damage are prevented or attenuated under such conditions. MATERIALS AND METHODS: Under methoxyflurane anesthesia, an enterectomy in the rat was performed by excision of the duodenum, jejunum, and ileum. The rats were then bled to and maintained at a mean arterial pressure of 40 mm Hg until 40% of the maximal shed volume was returned in the form of Ringer's lactate. The animals were then resuscitated with four times the volume of shed blood with Ringer's lactate over 1 h. At 1.5 h after the completion of resuscitation, hepatocellular function [i.e., the maximal velocity (Vmax) and transport efficiency (Km) of indocyanine green (ICG) clearance] was assessed by an in vivo ICG clearance technique. Blood samples were taken for the measurement of TNF, IL-6, and liver enzymes (i.e., SGPT and SGOT). Cardiac output and microvascular blood flow were determined by ICG dilution and laser Doppler flowmetry, respectively. RESULTS: The increase in circulating levels of TNF but not IL-6 was prevented by enterectomy prior to hemorrhage. The reduced Vmax and K(m) and elevated SGPT and SGOT following hemorrhage and resuscitation, however, were not significantly affected by prior enterectomy. Moreover, enterectomy before hemorrhage further reduced hepatic perfusion. CONCLUSION: Since enterectomy prior to the onset of hemorrhage does not prevent or attenuate the reduced ICG clearance and elevated liver enzymes despite downregulation of TNF production, it appears that the small intestine does not play a significant role in producing hepatocellular dysfunction and injury following trauma and hemorrhagic shock.

Alanine Transaminase↗

PubMed1998

Is Fas ligand or endotoxin responsible for mucosal lymphocyte apoptosis in sepsis?

BACKGROUND: Apoptosis (Ao) is a normal constitutive process that seems to have pathological effects in diseases of immune deficiency and autoimmune disorders, as well as in certain lymphoid tissues during sepsis. Little is known about this process in mucosal lymphoid tissue, such as intestinal intraepithelial lymphocytes (IELs). OBJECTIVES: To determine whether sepsis induces increased Ao in small intestinal IEL, whether this was associated with functional changes in cytokine gene expression in the IEL, and which mediators control this process and their impact on the survival of the mouse with sepsis. DESIGN: Male C3H/HeN (endotoxin-sensitive), C3H/HeJ (endotoxin-tolerant), and C3H/HeJ-FasLgld (endotoxin-tolerant/Fas ligand [FasL]-deficient) mice were subjected to sepsis (cecal ligation and puncture [CLP]) and IELs were harvested at 4 (early) or 24 hours (late sepsis). Alterations in the cell phenotype and Ao (TUNEL [terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick-end labeling] assay) were determined by 3-color flow cytometry. Cytokine gene expression was assessed by multiprobe RNase protection assay. RESULTS: At 4 hours after CLP, only the frequency of IEL which was CD8+ decreased markedly. By 24 hours after CLP, the number of CD8+ and CD4+ cells decreased while the proportion of double-negative cells showed a marked increase when compared with sham-controls. The percentage of Ao positive in CD8+ and CD4+ double-positive and double-negative cells increased markedly 24 hours after CLP concomitant with a significant (P<.05 vs sham-controls, Mann-Whitney U test) increase in expression of the IL-2, IL-10, and IL-15 gene. These data collectively suggest that sepsis causes lymphocyte activation-induced Ao that may be mediated by FasL. Additional studies were done to determine if the increased Ao was due to either endotoxin or FasL. The results of studies with endotoxin-tolerant C3H/HeJ or FasL-deficient C3H/HeJ-FasLgld mice showed an increase in A,, in CD4+ and CD8+ cells from septic C3H/HeJ but not C3H/HeJ-FasLgld mice. With regard to septic mortality, our results indicated that there was a marked reduction in mortality in C3H/HeJ-FasLgld vs C3H/HeJ mice. CONCLUSIONS: We conclude that the phenotypic changes associated with increased Ao may be a reflection of localized immune cell activation due to a FasL-mediated process and not endotoxin. Thus, FasL directly and/or indirectly contributes to higher septic mortality.

Animals↗

PubMed1998

Dehydroepiandrosterone: an inexpensive steroid hormone that decreases the mortality due to sepsis following trauma-induced hemorrhage.

BACKGROUND: Recent studies suggest that male sex steroids play a role in producing immunodepression following trauma-hemorrhage. This notion is supported by studies showing that castration of male mice before trauma-hemorrhage or the administration of the androgen receptor blocker flutamide following trauma-hemorrhage in noncastrated animals prevents immunodepression and improves the survival rate of animals subjected to subsequent sepsis. However, it remains unknown whether the most abundant steroid hormone, dehydroepiandrosterone (DHEA), protects or depresses immune functions following trauma-hemorrhage. In this regard, DHEA has been reported to have estrogenic and androgenic properties, depending on the hormonal milieu. OBJECTIVE: To determine whether administration of DHEA after trauma-hemorrhage has any salutary or deleterious effects on immune responses, and whether it improves the survival of animals subjected to subsequent sepsis. DESIGN: Male C3H/HeN mice underwent laparotomy (ie, trauma-induced) and hemorrhagic shock (blood pressure, 35+/-5 mm Hg for 90 minutes) followed by fluid resuscitation, or sham operation. The animals then received 100 mg of DHEA (4 mg/kg) or propylene glycol (hereafter referred to as vehicle). At 24 hours after trauma-hemorrhage and resuscitation, the animals were killed and blood, spleens, and peritoneal macrophages were harvested. Splenocyte proliferation and interleukin (IL) 2 release and splenic and peritoneal macrophage IL-1 and IL-6 release were determined. In a separate set of experiments, sepsis was induced by cecal ligation and puncture at 48 hours after trauma-hemorrhage and resuscitation. For those studies, the animals received vehicle, a single 100-microg dose of DHEA, or 100 microg/d DHEA for 3 days following hemorrhage and resuscitation. Survival was monitored for 10 days after the induction of sepsis. RESULTS: Administration of DHEA restored the depressed splenocyte and macrophage functions at 24 hours after trauma-hemorrhage. Moreover, daily administration of DHEA for 3 days significantly increased the survival of animals subjected to subsequent sepsis (P=.01). CONCLUSION: The finding that DHEA markedly improves the depressed immune functions and survival of animals subjected to subsequent sepsis suggests that short-term treatment with DHEA after trauma-hemorrhage is a safe and novel approach for preventing immunodepression and for decreasing the mortality rate due to subsequent sepsis.

Adjuvants, Immunologic↗

PubMed1998

The pivotal role of adrenomedullin in producing hyperdynamic circulation during the early stage of sepsis.

BACKGROUND: Initial cardiovascular responses during sepsis are characterized by hyperdynamic circulation. Although studies have shown that a novel potent vasodilatory peptide, adrenomedullin (ADM), is up-regulated under such conditions, it remains unknown whether ADM is responsible for initiating the hyperdynamic response. OBJECTIVE: To determine whether increased ADM release during early sepsis plays any major role in producing hyperdynamic circulation. DESIGN, INTERVENTION, AND MAIN OUTCOME MEASURE: Synthetic rat ADM (8.5 microg/kg of body weight) was infused intravenously in normal rats for 15 minutes at a constant rate. Cardiac output, stroke volume, and microvascular blood flow in various organs were determined immediately as well as 30 minutes after ADM infusion. At 30 minutes after infusion, plasma ADM level was also measured. In additional groups, rats were subjected to sepsis by cecal ligation and puncture. At 1.5 hours after cecal ligation and puncture, specific anti-rat ADM antibodies were infused, which completely neutralized the circulating ADM. Various hemodynamic variables were measured 5 hours after cecal ligation and puncture (ie, the early stage of sepsis). RESULTS: Cardiac output, stroke volume, and microvascular blood flow in the liver, small intestine, kidney, and spleen increased, and total peripheral resistance decreased 0 and 30 minutes after ADM infusion. In addition, plasma levels of ADM increased from the preinfusion level of 92.7+/-5.3 to 691.1+/-28.2 pg/mL 30 minutes after ADM infusion, which was similar to ADM levels observed during early sepsis. Moreover, 5 hours after the onset of sepsis, cardiac output, stroke volume, and microvascular blood flow in various organs increased and total peripheral resistance decreased. Administration of anti-ADM antibodies, however, prevented the occurrence of the hyperdynamic response. CONCLUSIONS: The results suggest that increased ADM production and/or release plays a major role in producing hyperdynamic responses during early sepsis. Since our previous studies have shown that vascular responsiveness to ADM decreases in late sepsis, maintenance of ADM vascular responsiveness by pharmacological agents during the course of sepsis may prevent transition from the hyperdynamic to the hypodynamic state.

Adrenomedullin↗

PubMed1998

Gut and liver: the organs responsible for increased nitric oxide production after trauma-hemorrhage and resuscitation.

OBJECTIVE: To determine which organs produce the increased levels of nitric oxide (NO) seen after hemorrhage and resuscitation. ANIMALS AND INTERVENTIONS: Adult male rats underwent laparotomy (ie, trauma induced) and were bled to and maintained at a mean arterial pressure of 40 mm Hg until 40% of the maximum bleedout volume was returned in the form of Ringer lactate. The rats were then resuscitated with Ringer lactate, 4 times the maximum bleedout volume for 1 hour. Sham-operated animals underwent only the surgical procedure. MAIN OUTCOME MEASURES: Plasma levels of nitrate/nitrite (NO3-/NO2-, stable products of NO) were measured by colorimetric assay at the maximum bleedout volume; at the end of hemorrhage; at the end of resuscitation; and 1.5, 4, 8, and 24 hours after resuscitation. In additional rats, the heart, liver, small intestine, kidneys, and spleen were harvested 4 hours after resuscitation for the measurement of NO3-/NO2- levels. Moreover, tissue perfusion was determined in the above-mentioned organs by radioactive microspheres 4 hours after resuscitation in other groups of animals. RESULTS: Plasma levels of NO3-/NO2- were similar to those of sham-operated animals during hemorrhage and at the end of resuscitation. One and a half hours after the end of resuscitation, however, NO production increased significantly. The peak levels of plasma NO3-/NO2- occurred at 4 hours, and the levels remained elevated even 24 hours after resuscitation. Tissue NO3-/NO2- levels were significantly increased in the liver, small intestine, and spleen 4 hours after resuscitation. In contrast, the levels of NO3-/NO2- were similar to those of sham-operated animals in the heart and kidneys at all times. Blood flow in the heart was maintained after hemorrhage, whereas hepatic, intestinal, splenic, and renal perfusion decreased significantly. CONCLUSIONS: The gut and liver seem to be the sites responsible for the increased NO production seen after trauma and hemorrhage. The overproduction of NO is most likely caused by up-regulation of inducible NO synthase. Thus, attempts to reduce NO production using specific inhibitors for inducible NO synthase might be helpful for improving hepatic and intestinal functions after trauma and hemorrhagic shock.

Animals↗

PubMed1998

Thermal injury alters macrophage responses to prostaglandin E2: contribution to the enhancement of inducible nitric oxide synthase activity.

Prostaglandin E2 (PGE2) and macrophage (Mphi)-derived reactive nitrogen intermediates (RNI) have been implicated in T cell dysfunction after thermal injury. Normally, Mphi inducible nitric oxide synthase (iNOS) activity can be regulated by PGE2, however, it is unknown whether PGE2 modulates Mphi iNOS activity after thermal injury. Splenic Mphi isolated from mice 7 days after thermal injury produced higher levels of RNI than Mphi from sham mice when stimulated with lipopolysaccharide (LPS) or interferon-gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) in combination. PGE2, when added concurrently with LPS, suppressed RNI production by Mphi from sham mice, whereas Mphi from injured mice were unaffected. When Mphi were pretreated with PGE2 before LPS, RNI production was suppressed in both populations. RNI production in response to IFN-gamma or IFN-gamma and TNF-alpha in combination was enhanced by PGE2 in both populations, however, the effect was markedly greater in Mphi from injured mice. The PGE2-mediated changes in RNI production were paralleled by similar changes in iNOS protein expression, suggesting that the effect of PGE2 was at the level of enzyme expression rather than activity. Dibutryl cAMP induced similar effects as PGE2, suggesting the response to PGE2 after thermal injury is independent of potential changes in PGE2-induced adenylate cyclase activity and is cAMP-mediated. The results indicate that Mphi from burned mice display an altered sensitivity to PGE2, resulting in enhanced iNOS activity. Thus, PGE2, which is elevated after thermal injury and can directly suppress T cell function, may also contribute to immune dysfunction through the enhancement of Mphi iNOS activity.

Animals↗

PubMed1998

Does hepatocellular injury in sepsis involve apoptosis?

Apoptosis (AO) is a process by which cells typically undergo a form of nonnecrotic cellular suicide. AO normally allows the host to selectively delete cells from a given tissue site without producing bystander injury associated with necrosis. However, inappropriate induction of AO has been associated with a variety of acute as well as chronic pathological states and may contribute to the therapeutic nonresponsiveness frequently encountered in the septic animal/patient's organ function. In this respect, while AO has been demonstrated in a variety of immune cell tissues of septic animals it is unclear if it is present in the septic liver. Therefore, it was the aim of our study to determine if AO is evident in hepatocytes of polymicrobial septic animals. To assess this, C3H/HeN male mice were subjected to polymicrobial sepsis (cecal ligation and puncture) or sham- CLP (Sham). Hepatocytes were then harvested at 4 h (early hyperdynamic phase) or 24 h (late hypodynamic state) later, and indices of AO were assessed [cell cycle analysis of Annexin V/propidium iodide (PI) staining for flow cytometric analysis, DNA extracted, and cell death ELISA]. Plasma glutamic pyruvic transaminase (GPT) was also colorimetrically assessed as well as total viable cell yield as an index of hepatocellular necrosis. The results indicate that indices of hepatocellular AO, as determined by cell cycle analysis and cell death ELISA, were markedly increased in polymicrobial septic mice at 24 h. However, while an increase in DNA fragmentation/degradation could be consistently detected, the pattern was typically faint. Similarly, although there was an increase in Annexin V staining it was not dissociated from that of PI (necrotic index). Alternatively, necrosis (as evidenced by increased GPT levels at both 4 and 24 h) preceded the induction of all the indices of AO. Taken together, these data suggest a role for both necrosis and apoptosis in the evolution of hepatocellular injury encountered in the polymicrobial septic animal/patient which may represent a unique pattern of cell death under such conditions.

Animals↗

PubMed1998

Sepsis induces increased apoptosis in lamina propria mononuclear cells which is associated with altered cytokine gene expression.

Studies indicate that lymphoid tissue (e.g., thymus, bone marrow, and Peyer's patches) shows evidence of increase apoptosis (Ao, a form of nonnecrotic cell death) during sepsis. However, it is not known if mucosal lymphoid tissue, such as lamina propria (LP), also shows evidence of increased Ao and if so, is this associated with functional changes, i.e., cytokine gene expression in the LP. To examine this, male C3H/HeN mice were subjected to cecal ligation and puncture (CLP) and lamina propria mononuclear cells (LPMC) were harvested at 4 h (early sepsis) or 24 h (late sepsis). Alterations in the cell phenotype as well as Ao (Tunel assay) were determined by three-color flow cytometry. Cytokine gene expression was assessed by multiprobe RNase protection assay. Sham LPMC preparations were found to be 34.4 +/- 2.4% B220(+) (B-cells), while 12.4 +/- 2.1% were CD8(+) (cytotoxic T-cells), 22.0 +/- 0.8% were CD4(+) (helper T-cells), and 6.4 +/- 0.7% were F4/80(+) (macrophages). The frequency of B220(+) (9%* upward arrow) and CD8 (6%* upward arrow) populations increased markedly at 4 h after CLP; however, this increase was not seen at 24 h. The percentage of Ao+ in CD8(+), B220(+), and F4/80(+) cells increased markedly at both 4 and 24 h. CD4(+) cells showed a marked increase in Ao only at 24 h after CLP. When LPMC mRNA expression was examined, a significant increase in IL-2, -10, and -15 gene expression was observed only at 24 h but not 4 h after CLP. Thus, the early phenotypic changes associated with increased Ao may be a reflection of localized immune cell activation in early sepsis contributing to the increased cytokine gene expression seen in late sepsis. This localized activation may contribute to gastrointestinal inflammation and/or immune dysfunction in sepsis.

Animals↗

PubMed1998

Does neutrophil-mediated oxidative stress play any significant role in producing hepatocellular dysfunction during early sepsis?

BACKGROUND: Although previous studies have indicated that hepatocellular dysfunction during early sepsis can be prevented by prior neutrophil depletion, it remains unknown whether the changes in hepatic oxidative stress induced by activated neutrophils are responsible for the salutary effect of neutropenia on hepatocellular function. MATERIALS AND METHODS: Neutropenia was induced in the rat by intravenous injection of immunoglobulins directly against rat neutrophils (anti-neutrophil Ig) at 16 and 2 h prior to the initiation of cecal ligation and puncture (CLP, i.e., an animal model of polymicrobial sepsis) or sham operation. Neutropenia was confirmed by peripheral blood smears. Neutrophil-competent control animals were given nonimmunized Ig prior to the onset of sepsis. Sham animals received anti-neutrophil Ig or control Ig. The levels of reduced glutathione (GSH) and oxidized glutathione (GSSG) in the liver were determined at 5 h after CLP (i.e., the early, hyperdynamic stage of sepsis) or sham operation by high performance liquid chromatography. RESULTS: Although the levels of hepatic GSH and GSSG decreased significantly at 5 h after CLP, irrespective of neutropenia, the ratio of GSSG/GSH was not significantly altered under such conditions. The decrease in hepatic glutathione concentrations in septic animals may represent the decreased synthesis or increased efflux into the bile or circulation. CONCLUSION: Since neutrophil depletion did not significantly affect hepatic levels of GSH and GSSG as well as the GSSG/GSH ratio but prevented the occurrence of hepatocellular dysfunction, factors other than oxidative stress are likely to be the mechanism responsible for depressing hepatocellular function during the early, hyperdynamic stage of sepsis.

Animals↗

PubMed1998

Increased gut permeability after hemorrhage is associated with upregulation of local and systemic IL-6.

Although intestinal barrier failure after hemorrhage is a well-documented event, the underlying mechanism is poorly understood. The aim of this study, therefore, was to determine whether altered intestinal permeability after hemorrhage is associated with upregulation of local and systemic interleukin-6 (IL-6). To study this, rats underwent laparotomy (i.e., trauma induced) and were bled to and maintained at a mean arterial pressure of 40 mm Hg until 40% of the shed blood volume was returned in the form of Ringer's lactate. The animals were then resuscitated with four times the volume of shed blood with Ringer's lactate over 60 min. At 1.5 h postresuscitation, an in vivo ligated loop of a distal small intestine was formed and the passage of 4-kDa fluorescein isothiocyanate-conjugated dextran (FD4) from the intestinal lumen into the portal vein and carotid artery blood was analyzed by fluorescence spectrometry. Samples from the portal vein and a carotid artery were collected and plasma IL-6 was assayed. Intraepithelial lymphocytes from a distal small intestine were isolated and cultured in vitro for 24 h with or without anti-rat CD3 monoclonal antibody stimulation. IL-6 activity in freshly isolated cells and its release by cultured lymphocytes were determined. Intestinal perfusion and portal blood flow were determined by radioactive microspheres in another set of parallel experiments. The results indicate that lumen-to-blood passage of FD4 through the wall of the small intestine increased significantly at 1.5 h after hemorrhage and resuscitation and was associated with decreased intestinal perfusion and portal blood flow. Plasma IL-6 levels in the portal vein and carotid artery markedly increased at 1.5 h after hemorrhage and resuscitation. In addition, a significant correlation was observed between plasma IL-6 and FD4 concentrations. Higher IL-6 activity in freshly isolated cells was found in hemorrhaged rats. Increased IL-6 release by cultured lymphocytes was also observed either with or without anti-rat CD3 monoclonal antibody stimulation. Thus, the increased intestinal permeability following trauma-hemorrhage and resuscitation appears to be associated with systemic and intestinal IL-6 upregulation.

Animals↗

PubMed1998

Hepatocellular dysfunction after severe hypotension in the absence of blood loss is associated with the increased IL-6 and PGE2.

Although hepatocellular dysfunction occurs following trauma and hemorrhagic shock, whether severe hypotension even in the absence of blood loss depresses hepatocellular function remains unknown. The aim of this study, therefore, was to determine whether chemically induced severe hypotension causes hepatocellular dysfunction and, if so, whether IL-6 and PGE2 are associated with this dysfunction. To study this, hypotension was induced in adult male rats by intravenous infusion of a high dosage of ATP-MgCl2 solution (3.2 +/- 0.45 micromol/min/kg body wt) for 60 min. Blood pressure decreased from 108 +/- 6 mm Hg to an average of 43 mm Hg during the infusion period and returned to normal levels immediately after the completion of ATP-MgCl2 infusion. At 0 and 4 h after hypotension, hepatocellular function [i.e., maximum velocity of indocyanine green clearance (Vmax) and its efficiency (Km)] was measured using a fiberoptic catheter and in vivo hemoreflectometer. Cardiac output was determined by dye dilution. Microvascular blood flow was assessed by laser Doppler flowmetry. Plasma levels of PGE2 and IL-6 were measured by radioimmunoassay and bioassay, respectively. The results indicate that severe hypotension in the absence of any blood loss depresses hepatocellular function (i.e., decreased Vmax and Km) despite stable cardiac output and hepatic perfusion at 0 and 4 h after the completion of hypotension. Moreover, severe hypotension resulted in significantly increased plasma levels of PGE2 (only at 0 h) and IL-6. Thus, chemically induced severe hypotension in the absence of any blood loss, which does not significantly reduce cardiac output and hepatic perfusion, depresses hepatocellular function and upregulates IL-6 and PGE2 production.

Adenosine Triphosphate↗

PubMed1998

[Contribution of soft tissue trauma and/or bone fracture to immune suppression after hemorrhagic shock in the animal experiment].

Bone fracture, soft-tissue trauma and hemorrhagic shock are frequent complications in trauma patients, and these patients are known to be immunocompromised. Nonetheless, it is difficult to differentiate the effect of soft-tissue trauma plus hemorrhage from that of bone fracture and hemorrhage on host immune function in the clinical setting. To determine this experimentally, closed bone fracture (right lower leg) and/or soft-tissue trauma (2.5 cm midline laparotomy) were induced prior to hemorrhagic shock (mean arterial BP of 35 +/- 5 mm Hg for 90 min) in male C3H/HeN mice. All animals were killed at 72 h after initiation of the experiment and the spleens were collected aseptically. More significant depression of splenocyte IL-2 and IL-3 release occurred with the combined insult than after bony injury or tissue trauma alone with hemorrhage. The present study suggests that different traumatic insults, i.e. bone fracture as well as soft-tissue trauma in conjunction with hemorrhagic shock, produce comparable depression of host immune function. Moreover, combination of closed bone fracture and soft-tissue trauma prior to hemorrhagic shock leads to even more compromised immunity. This indicates that different mechanisms of immune depression may be involved following soft-tissue trauma or bony injury coupled with hemorrhage. The markedly depressed immune function following bony injury, soft-tissue trauma and hemorrhagic shock may contribute to the increased susceptibility of severely injured patients to sepsis and the ensuing multiple organ failure in the clinical situation.

Animals↗