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T G Buchman

Publications and source records attributed to T G Buchman.

At least 73 records · Page 4Linked to original sources
PubMed1995

Molecular biology in the intensive care unit: a framework for interpretation.

The tools of cellular and molecular biology provide insight into the mechanisms operating in cellular functions and have already advanced the diagnosis and treatment of human disease. The goal of this article is to provide a framework utilizing recent advances in basic science. Changing the frame of analysis from clinically familiar systems physiology to the interior of a cell, and then considering the cell as an information repository and processor, may simplify the interpretation of molecular and cellular studies. Gene therapy and manipulation of signal transduction pathways are used to demonstrate how the framework can be applied to augment clinicians' understanding of these processes.

Critical Care↗
PubMed1995

Endotoxin induces organ-specific endothelial cell injury.

Endothelial cell (EC) injury is observed in clinically important pathological processes, including bacterial endotoxemia. We hypothesized that such pathological processes may exhibit target organ heterogeneity due to organ-specific heterogeneity of endothelial cells. To test this hypothesis, endothelial cells of aorta (AO), pulmonary artery (PA), left ventricle (LV), and right ventricle (RV) were cultured from individual sheep and exposed to bacterial endotoxin. Marked heterogeneity in endotoxin-induced cytotoxicity was observed. AOEC were the most sensitive, followed by PAEC, LVEC, and RVEC. This cytotoxicity was manifested as programmed cell death (apoptosis). All cells were able to express both interleukin-6 and endothelin-1 (ET-1) transcripts. Following exposure to bacterial endotoxin, interleukin-6 transcripts accumulated in all cells, whereas ET-1 expression was constant or slightly decreased. These data suggest that organ-specific heterogeneity of EC responsiveness to endotoxin is a potential determinant of organ-specific resistance to endotoxin and other mediators of injury.

Animals↗
PubMed1994

Antioxidants modulate induction of programmed endothelial cell death (apoptosis) by endotoxin.

OBJECTIVE: To evaluate the potential role of reactive oxygen metabolites as signals for endothelial cell apoptosis. DESIGN: A series of antioxidants were evaluated for their ability to block apoptosis in cultured porcine aortic endothelial cells in vitro. RESULTS: Scavenging of the hydroxyl radical with the membrane-permeable scavenger dimethyl sulfoxide or blocking its generation via the Fenton reaction by the chelation of iron with o-phenanthroline blocked apoptosis, whereas the cell membrane-impermeable scavengers superoxide dismutase and catalase did not block apoptosis. Inhibition of xanthine oxidase with enzyme-inhibitory levels of allopurinol also failed to block apoptosis, whereas high levels of allopurinol, which also scavenge the hydroxyl radical in vitro, conferred protection. In each case (dimethyl sulfoxide, o-phenanthroline, and high-dose allopurinol), hydroxyl radical ablation was only effective when administered before the priming step (lipopolysaccharide) and was ineffective when administered later, prior to the activation step (heat shock). CONCLUSIONS: These findings suggest a novel role for the hydroxyl radical as a nonlethal intracellular signal in endothelial cell apoptosis. Moreover, the results support a role for programmed cell death in the pathogenesis of multiple organ dysfunction syndrome and suggest novel strategies for prophylaxis and therapy of the most common cause of death in surgical intensive care units.

Allopurinol↗
PubMed1994

Induction of heat-shock gene expression in postischemic pig liver depends on superoxide generation.

BACKGROUND/AIMS: Both hemorrhagic and cardiogenic shock are associated with hepatic shock gene expression at resuscitation. This study investigated the potential role of intravascular superoxide anion as a proximal trigger of heat shock protein gene expression. METHODS: Preanesthetized pigs were subjected to 120 m of total warm hepatic ischemia. The survival model consisted of warm, total hepatic ischemia and reperfusion (with active portal-systemic bypass) followed by reperfusion and survival for 3 days. Serial hepatic biopsy samples were evaluated for the expression of heat shock protein 72 (HSP-72) messenger RNA (mRNA) by Northern and Western analysis and by in situ RNA hybridization. The possible role of intravascular O2- as a mediator of heat shock response was evaluated by its specific inhibition by the intravenous infusion of recombinant human superoxide dismutase (SOD). RESULTS: Ischemia for 120 minutes followed by 60 minutes of reperfusion caused accumulation of HSP-72 mRNA. Transcripts were localized to hepatocytes. HSP-72 mRNA was detected neither following ischemia alone nor when SOD was infused for 15 minutes at reperfusion. Three days later, transcripts were not detectable, but HSP-72 protein accumulated irrespective of SOD administration. CONCLUSIONS: Warm hepatic ischemia induces the hepatocyte expression of HSP-72 at reperfusion by a mechanism that is dependent upon the superoxide anion, probably generated intravascularly. However, the transient dismutation of superoxide is insufficient to suppress subsequent accumulation of HSP-72.

Animals↗
PubMed1994

A comparison of statistical and connectionist models for the prediction of chronicity in a surgical intensive care unit.

OBJECTIVE: To compare statistical and connectionist models for the prediction of chronicity which is influenced by patient disease and external factors. DESIGN: Retrospective development of predictive criteria and subsequent prospective testing of the same predictive criteria, using multiple logistic regression and three architecturally distinct neural networks; revision of predictive criteria. SETTING: Surgical intensive care unit (ICU) equipped with a clinical information system in a +/- 1000-bed university hospital. PATIENTS: Four hundred ninety-one patients with ICU length of stay 3 days who survived at least an additional 4 days. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Chronicity was defined as a length of stay > 7 days. Neural networks predicted chronicity more reliably than the statistical model regardless of the former's architecture. However, the neural networks' ability to predict this chronicity degraded over time. CONCLUSIONS: Connectionist models may contribute to the prediction of clinical trajectory, including outcome and resource utilization, in surgical ICUs.

Evaluation Studies as Topic↗
PubMed1994

Limited role of barium enema examination preceding colostomy closure in trauma patients.

Whether trauma patients should undergo barium enema (BE) examination of the colon prior to colostomy closure has recently been questioned. To ascertain the utility of BE and its impact on postoperative course in this patient population, we reviewed 86 trauma patients who underwent colostomy closure during a 12-year period at our institution. There were 82 males and four females with an average age of 28 years. Ninety-five percent of the injuries were the result of penetrating trauma. Sixteen patients had rectal injuries. Fifteen of these had BE greater than 6 weeks post-trauma and all showed healing of the injury. Of the 70 patients with colonic injuries, 43 (group 1) had BE prior to colostomy closure. Ninety-eight percent (n = 42) of these studies were negative. The only positive finding did not affect the planned surgical procedure. Group 2 (n = 27) did not have a BE prior to colostomy closure. Overall complication rates were not significantly different between group 1 (18.6%) and group 2 (29.6%). We conclude that BE prior to colostomy closure for colonic injuries yields little useful information and does not affect the morbidity rate prior to colostomy closure. Its routine usage should be abandoned. The role of barium enema in assessing rectal injury status is less clear because of the small number in our series, but probably offers no advantage over proctoscopy.

Adolescent↗
PubMed1994

Myocardial heat shock gene expression in pigs is dependent on superoxide anion generated at reperfusion.

The heat shock response is a conserved response to cell injury. We sought to determine if ischemia alone versus events at reperfusion stimulated expression of the major heat shock protein (hsp-72) in a clinically relevant model of global myocardial ischemia in pigs. Pigs were placed on nonpulsatile cardiopulmonary bypass. Serial transmural cardiac biopsies were taken at baseline following 20 min of normothermic global ischemia (induced by crossclamping the aorta) and at 20, 40, and 60 min of reperfusion. Test animals received a bolus and subsequent aortic root infusion of superoxide dismutase (total 7,500 U/kg) beginning just prior to reperfusion. Hsp-72 mRNA abundance was estimated from Northern blots. We found that hsp-72 mRNA was not induced following 20 min of ischemia but accumulated to high levels within 20 min of reperfusion. Intravascular administration of superoxide dismutase at reperfusion eliminated hsp-72 mRNA induction. We conclude that in the postischemic myocardium, hsp-72 gene expression is dependent on superoxide anion generation at reperfusion. In this setting, hsp-72 gene expression may reflect a specific response to oxidative injury rather than a more general response to metabolic stress associated with ischemia.

Adenosine Triphosphate↗
PubMed1994

Thiol reducing agents modulate induced apoptosis in porcine endothelial cells.

When cultured porcine endothelial cells are exposed first to endotoxin (lipopolysaccharide (LPS)) followed by standard inducers of the heat shock response in vitro (heat or sodium arsenite), these cells aberrantly execute programmed cell death. This cell death is dependent upon two distinct events: the LPS-priming step and the heat shock-induced activation step. Prior work demonstrated that the LPS-priming step could be blocked by the prior application of cell-permeable hydroxyl radical scavengers, suggesting a role for this reactive oxygen species as an important intracellular signal mediating the first step. In these present experiments, we evaluated the potential role of reduction-oxidation mechanisms in the heat shock activation step. The thiol reducing agents reduced glutathione (GSH), n-acetylcysteine (NAC), and dithiothreitol (DTT) were evaluated for their ability to block programmed cell death in LPS-primed porcine aortic endothelial cells. Both DTT and NAC, agents that augment intracellular reduced glutathione levels, were protective against cell death when applied prior to heat shock induction with sodium arsenite (As) in endothelial cells treated previously with LPS. The less cell permeable agent GSH was not protective. Delayed application of DTT or NAC could block progression to cell death for up to 1.5 h after initiation of the heat shock response with As. These data show that heat shock-induced programmed cell death in LPS-primed endothelial cells can be arrested, at least in its early stages, by agents that augment or stabilize the reducing potential of the cell.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcysteine↗
PubMed1994

Heat shock-induced cell death in murine microvascular endothelial cells depends on priming with tumor necrosis factor-alpha or interferon-gamma.

We previously reported that sequential exposure of cultured porcine aortic endothelial cells to lipopolysaccharide (LPS) and then to inducers of the heat shock response resulted in lethal injury by programmed cell death. In these present experiments, we evaluated the ability of other mediators of sepsis to prime endothelial cells for subsequent injury upon heat shock induction. We used SVEC4-10 microvascular endothelial cells, a cloned SV40-transformed cell line derived from LPS-resistant C3H/HeJ mice. When these endothelial cells were treated first with tumor necrosis factor-alpha followed by induction of the heat shock response with either heat or sodium arsenite (As), a standard chemical inducer of the heat shock response in vitro, a tumor necrosis factor dose-dependent cytotoxicity was observed, similar to that which we had seen previously in porcine endothelial cells primed with LPS. Subsequent experiments found that priming with interferon-gamma produced a similar dose-dependent toxicity upon heat shock with either heat or As. The reducing agents dithiothreitol and n-acetylcysteine, which we have previously shown to inhibit heat shock-induced programmed cell death in LPS-primed porcine endothelial cells, were also effective in protecting against heat shock-induced death following cytokine-priming in this microvascular cell line, suggesting that the intracellular signaling pathways of these priming agents are somewhere convergent. These data suggest that both exogenous and endogenous mediators of inflammation can prime endothelial cells for subsequent injury upon induction of the heat shock response, and are consistent with the emerging concept of redundancy in inflammatory signaling.

Acetylcysteine↗
PubMed1994

Rapid detection and identification of Candida albicans and Torulopsis (Candida) glabrata in clinical specimens by species-specific nested PCR amplification of a cytochrome P-450 lanosterol-alpha-demethylase (L1A1) gene fragment.

PCR of a Candida albicans cytochrome P-450 lanosterol-alpha-demethylase (P450-L1A1) gene segment is a rapid and sensitive method of detection in clinical specimens. This enzyme is a target for azole antifungal action. In order to directly detect and identify the clinically most important species of Candida, we cloned and sequenced 1.3-kbp fragments of the cytochrome P450-L1A1 genes from Torulopsis (Candida) glabrata and from Candida krusei. These segments were compared with the published sequences from C. albicans and Candida tropicalis. Amplimers for gene sequences highly conserved throughout the fungal kingdom were first used; positive PCR results were obtained for C. albicans, T. glabrata, C. krusei, Candida parapsilosis, C. tropicalis, Cryptococcus neoformans, and Trichosporon beigelii DNA extracts. Primers were then selected for a highly variable region of the gene, allowing the species-specific detection from purified DNA of C. albicans, T. glabrata, C. krusei, and C. tropicalis. The assay sensitivity as tested for C. albicans in seeded clinical specimens such as blood, peritoneal fluid, or urine was 10 to 20 cells per 0.1 ml. Compared with results obtained by culture, the sensitivity, specificity, and efficiency of the species-specific nested PCR tested with 80 clinical specimens were 71, 95, and 83% for C. albicans and 100, 97, and 98% for T. glabrata, respectively.

Amino Acid Sequence↗
PubMed1993

Induction of translational thermotolerance in liver of thermally stressed rats.

Heat-shock gene expression in cultures of single cell types has been well characterized but little is known about the heat-shock response of intact organs in vivo. In this study, the kinetics of hepatic heat-shock gene expression and the induction of thermotolerance were characterized in rats. Animals were subjected to a defined, reversible stress by increasing the core body temperature to 41 degrees C or 42 degrees C for 30 min. New synthesis of the inducible form of the heat shock-70 family of proteins (hsp-72) peaked simultaneously with the maximal level of hsp-72 transcripts at both temperatures. These data are consistent with previous observations in cultures of hepatoblastoma cells after thermal stress [De Maio, A., Beck, S. C. & Buchman, T. G. (1993) Circ. Shock 40, 177-186]. The incorporation of radioactive amino acids into polypeptides by the liver was blocked during the first hour of recovery after heat shock at 42 degrees C. This inhibition of protein synthesis by thermal stress could be prevented by prestressing rats at 42 degrees C for 30 min and allowing the rats to recover for 24 h at normal body temperature (37 degrees C). This phenomenon, previously defined as 'translational thermotolerance', correlates with the hepatic content of hsp-72; maximal protection occurs 24 h after a 42 degrees C thermal stress when hsp-72 (protein) is also maximum and decreases with the clearance of hsp-72 from the liver. These data suggest that the presence of hsp-72 within the liver may modulate the organ response to subsequent stresses and may be important to organ and animal survival after repeated insults.

Adaptation, Physiological↗
PubMed1993

Morbidity and timing of colostomy closure in trauma patients.

The reported morbidity of colostomy closure in trauma patients varies from 5 to 27 per cent. Low morbidity rates are cited as a factor favouring colostomy creation and against expanded indications for primary repair in the treatment of colonic injuries. In order to assess the morbidity of colostomy closure, we reviewed all colonic injuries from 1979 to 1991 at our institutions. In all, 86 trauma patients who underwent colostomy creation and closure were identified. There were 82 men and four women with an age range of 16 to 74 years (mean 28.1 years). Of these, 95 per cent (N = 82) resulted from penetrating trauma. Of the patients, 63 per cent (N = 54) received end colostomies and 81 per cent (N = 70) of the patients had associated injuries. Of the patients, 38 per cent (N = 33) had a complication with their initial operation. There were no deaths after colostomy closure, but a total morbidity of 24.4 per cent (N = 21) was noted. There were 11 anastomotic complications (two of which required repeat laparotomy) and nine wound infections. The average length of stay was 10.4 days. Morbidity was concentrated in the group who had complications at their initial hospitalization. This was especially true if these patients underwent closure earlier than 3 months after injury. Conversely, if the first operation was uncomplicated, waiting longer than 3 months to perform colostomy closure did not improve results further.

Adolescent↗
PubMed1993

Regulation of acute phase gene expression following surgery and endotoxin administration in the anesthetized pig.

BACKGROUND: The hepatic acute phase response (APR) reflects an organism's integrated response to stress. This APR results in augmented synthesis and secretion of specific procoagulants and antiproteases and a complementary decrease in the synthesis and secretion of several constitutive proteins, such as albumin. The cytokines tumor necrosis factor (TNF) or interleukin-6 (IL-6) have been identified as proximal mediators of the APR in response to endotoxin stress. The authors hypothesized that TNF, IL-6, or both would be the proximal mediators of the APR in response to anesthesia and surgical stress. METHODS: The effects of a standardized surgical stress on the APR in pigs under general anesthesia with sodium pentobarbital and ketamine hydrochloride was investigated. Acute phase gene transcription was assayed in nuclei from serial liver biopsies obtained before and after 2.5 h of surgical stress, and after endotoxin administration. Tumor necrosis factor and IL-6 mRNA levels in this liver tissue were examined by Northern blot hybridization, and simultaneous plasma levels of these cytokines were measured using bioassays. RESULTS: The transcription rates of three positive acute phase genes--chymotrypsin inhibitor, inter-alpha-trypsin inhibitor and beta-fibrinogen--increased seven-, six-, and twofold, respectively (P < 0.05), and the transcription rate of albumin, a negative acute phase gene, decreased to 34% of baseline (P < 0.01) during the 2.5 h of anesthesia and surgical stress. During this initial 2.5 h, plasma concentrations of TNF and IL-6 did not change. Hepatic IL-6 mRNA expression was never observed, and TNF mRNA expression was undetectable in six of seven pigs. Subsequent 10-micrograms/kg endotoxin administration caused 20- and 100-fold increases in plasma concentrations of TNF and IL-6, respectively (P < 0.01), and were associated with substantial hepatic expression of the TNF and IL-6 mRNAs. These increments in cytokines were not associated with any further increase in the acute phase gene transcription rates. Thus, the APR was initially regulated at the transcriptional level during surgical stress independent of, and not augmentable by, an endotoxin-provoked increase in either plasma levels or hepatic mRNA expression of TNF or IL-6. CONCLUSIONS: Surgical stress induced hepatic acute phase gene transcription within 2.5 h in the absence of either systemic or local (hepatic) increases in TNF or IL-6. Subsequent endotoxin-induced increases in TNF or IL-6 did not alter this surgical stress-induced acute phase gene transcription.

Acute-Phase Proteins↗
PubMed1993

Optimizing prehospital triage criteria for trauma team alerts.

Although scores and other prehospital triage schema effectively identify injured patients who will benefit from trauma center care, those tools are relatively nonspecific. One consequence is overtriage--transport of less severely injured patients to trauma centers--with resulting expenditure of scarce resources on patients who do not benefit from an emergent and intensive response. We developed a tool that, during the prehospital phase, can sort inner-city trauma victims into those who will require ICU/OR services and those who will not. Beginning with the trauma triage criteria promulgated by the American College of Surgeons, we created two tiers of personnel response based on vital signs, mechanism of injury, and limited anatomic information ordinarily reported by prehospital providers. Based on our initial experience with the two-tier response, the sorting criteria were revised and refined. The sensitivity of the current version of the two-tier criteria for predicting which trauma patients will require ICU/OR services during the first 24 hours of hospitalization approaches 95% (excluding misapplications of the tool) while avoiding urgent trauma team mobilization in 57% of patients triaged to our trauma center. Two-tiered trauma responses appear to be safe and may represent an important strategy for more effective distribution of increasingly scarce and costly resources.

Adult↗
PubMed1993

Induction of heat shock response leads to apoptosis in endothelial cells previously exposed to endotoxin.

The homeostatic response of complex eukaryotes to the challenge of environmental stress includes the induction of several programs of gene expression; among them are those for the acute phase genes and those for the heat shock genes. In some systems, the heat shock response, which is often elicited by more severe stimuli, preempts the acute phase response, which is seen in response to less severe challenges, as well as constitutive gene expression. Nevertheless, each response appears to provide a natural selective advantage for survival of the organism in a toxic environment. However, when cultured porcine endothelial cells were exposed first to a nonlethal level of bacterial endotoxin lipopolysaccharide (LPS), an inducer of the acute phase response, and then, simultaneously to standard stimuli, which normally elicit a salutary heat shock response, the cells died manifesting a pattern of DNA fragmentation (nucleolysis) characteristic of programmed cell death (apoptosis). The treatment of LPS-exposed cells with cycloheximide to block protein synthesis reproduced the lethal apoptosis that had been elicited by the induction of heat shock gene expression. Therefore, the preemption of other programs of stress gene expression by the prioritized expression of heat shock genes is associated with apoptosis.

Animals↗
PubMed1993

A reversibly palmitoylated resident protein (p63) of an ER-Golgi intermediate compartment is related to a circulatory shock resuscitation protein.

The recently identified 63 kDa membrane protein, p63, is a resident protein of a membrane network interposed in between rough ER and Golgi apparatus. To characterize p63 at the molecular level a 2.91 kb cDNA encoding p63 has been isolated from a human placenta lambda gt10 cDNA library. Sequence analysis of tryptic peptides prepared from isolated p63 confirmed the identify of the cloned gene. The translated amino acid sequence consists of 601 amino acids (65.8 kDa) with a single putative membrane-spanning region and a N-terminal cytoplasmic domain of 106 amino acids. The human p63 cDNA exhibits a high level of sequence identify to the pig hepatic cDNA 3AL (accession number M27092) whose expression is enhanced after resuscitation from circulatory shock. An additional remarkable feature of p63 is that it becomes reversibly palmitoylated when intracellular protein transport is blocked by the drug brefeldin A. Overexpression of p63 in COS cells led to the development of a striking tubular membrane network in the cytoplasm. This suggests that the protein may be determinant for the structure of the p63 compartment.

Amino Acid Sequence↗