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Biomedical subjects

M S Pasternack

Publications and source records attributed to M S Pasternack.

At least 19 recordsLinked to original sources
PubMed2001

Antibiotic prophylaxis for group A streptococcal burn wound infection is not necessary.

BACKGROUND: Historically, group A beta-hemolytic streptococci (GAS) burn wound infection has been a major source of morbidity and mortality in burn patients and has prompted the prophylactic administration of antibiotics to children with burns. Wound monitoring, surveillance cultures, and early excision of deep wounds may have changed this. Our objective in this project was to determine the efficacy of routine antibiotic prophylaxis in the era of early excision and closure of deep burn wounds. METHODS: Two cohorts of burned children were compared: all children admitted during calendar years 1992 through 1994 (group 1) and during calendar years 1995 through 1997 (group 2). All group 1 children received routine GAS antibiotic prophylaxis. Only those group 2 children with documented positive admission or surveillance cultures for GAS were treated. RESULTS: There were 511 children in group 1 and 406 children in group 2. They were well matched for age (4.7 +/- 0.21 years vs. 5.3 +/- 0.26 years, p = 0.06) and burn size (11.0% +/- 0.7% vs. 12.4% +/- 0.8%, p = 0.18). GAS species were recovered at admission or during hospitalization from 11 (2.6%) of group 1 children and 18 (4.4%) of group 2 children (p = 0.05), indicating a marginally higher rate of carriage in group 2. Nevertheless, in group 1 there were three (0.6%) who developed GAS wound infection and in group 2 there were four (0.98%, p = 0.71). The incidence of GAS infection in those patients with positive admission cultures was three (27%) of group 1 and four (22%) of group 2. No child developed fulminant GAS infection. CONCLUSION: Routine antibiotic prophylaxis of burn wounds in children in not effective in further reducing a low baseline incidence of GAS wound infection if admission screening by culture is used to identify those children who carry the organism and early excision of deep burns is practiced.

Administration, Oral↗
PubMed2000

Induction of rapid histone degradation by the cytotoxic T lymphocyte protease Granzyme A.

The cytotoxic T lymphocyte protease granzyme A induces caspase-independent cell death in which DNA single-strand nicking is observed instead of oligonucleosomal fragmentation. Granzyme A is a specific tryptase that concentrates in the nucleus of targeted cells and synergistically enhances DNA fragmentation induced by the caspase activator granzyme B. Here we show that granzyme A treatment of isolated nuclei enhances DNA accessibility to exogenous endonucleases. In vitro and after cell loading with perforin, GrnA completely degrades histone H1 and cleaves core histones into approximately 16-kDa fragments. Histone digestion provides a mechanism for unfolding compacted chromatin and facilitating endogenous DNase access to DNA during T cell and natural killer cell granule-mediated apoptosis.

Animals↗
PubMed2000

Cloning and expression of secretagogin, a novel neuroendocrine- and pancreatic islet of Langerhans-specific Ca2+-binding protein.

We have cloned a novel pancreatic beta cell and neuroendocrine cell-specific calcium-binding protein termed secretagogin. The cDNA obtained by immunoscreening a human pancreatic cDNA library using the recently described murine monoclonal antibody D24 contains an open reading frame of 828 base pairs. This codes for a cytoplasmic protein with six putative EF finger hand calcium-binding motifs. The gene could be localized to chromosome 6 by alignment with GenBank genomic sequence data. Northern blot analysis demonstrated abundant expression of this protein in the pancreas and to a lesser extent in the thyroid, adrenal medulla, and cortex. In addition it was expressed in scant quantity in the gastrointestinal tract (stomach, small intestine, and colon). Thyroid tissue expression of secretagogin was restricted to C-cells. Using a sandwich capture enzyme-linked immunosorbent assay with a detection limit of 6.5 pg/ml, considerable amounts of constitutively secreted protein could be measured in tissue culture supernatants of stably transfected RIN-5F and dog insulinoma (INS-H1) cell clones; however, in stably transfected Jurkat cells, the protein was only secreted upon CD3 stimulation. Functional analysis of transfected cell lines expressing secretagogin revealed an influence on calcium flux and cell proliferation. In RIN-5F cells, the antiproliferative effect is possibly due to secretagogin-triggered down-regulation of substance P transcription.

Amino Acid Sequence↗
PubMed2000

Cutaneous herpetic infections complicating burns.

Many people harbor herpes simplex virus, often with a known history of "cold sores". During the relatively immunosuppressed state associated with a serious burn, recrudescence of such infections can occur. We report four adults and two children who developed severe herpetic ulceration, over the face and neck in five patients and in a partial thickness wound in one patient. Herpetic infection was diagnosed by culture and direct immunofluorescence testing and treatment was immediately instituted with systemic and topical Acylovir(R) (Zovirax, Glaxo Wellcome). Ulceration healed under treatment and did not leave visible scarring in any of the patients. Although these infections are rapidly progressive, they respond to prompt treatment with antiviral chemotherapy. Rapidly progressive vesicles and ulceration appearing on the face or in the wounds of burn patients should prompt immediate evaluation for herpetic infection.

Acyclovir↗
PubMed1999

Inhibitory effect of haptoglobin on granulocyte chemotaxis, phagocytosis and bactericidal activity.

Human haptoglobin (Hp) is synthesized at hepatic and extrahepatic sites as an acute-phase reactant protein (APP). We investigated the effects of Hp on granulocyte function. The chemotaxis of freshly isolated human granulocytes and differentiated HL-60 cells in response to the bacterial tripeptide, f-met-leu-phe, was inhibited in the presence of a physiological concentration of Hp, but chemotaxis in the presence of the proinflammatory cytokine interleukin-8 (IL-8) was not inhibited. Phagocytosis of viable Escherichia coli, as well as fluoresceinated nonviable E. coli, was inhibited. Hp also reduced granulocyte intracellular bactericidal activity against E. coli. The observed inhibitory effects of Hp on granulocyte function are similar to those reported for C-reactive protein and suggest that APPs dampen the acute inflammatory response.

Blood Bactericidal Activity↗
PubMed1998

Beta-chemokines are released from HIV-1-specific cytolytic T-cell granules complexed to proteoglycans.

CD8+ lymphocytes are believed to be important in host defence against the human immunodeficiency virus (HIV)-1, inhibiting HIV-1 replication through both cytolytic and non-cytolytic pathways. The cytolytic pathway involves calcium-dependent exocytosis of perforin and granzyme proteases, as well as Fas-mediated programmed cell death, whereas the noncytolytic pathway involves the release of chemokines that prevent viral entry. Using granzyme A as a marker of cytolytic granule proteins, and macrophage inflammatory protein (MIP)-1alpha and RANTES as markers of HIV-1 inhibitory chemokines, we show that these two very different mediators of viral inhibition are both localized in the cytolytic granules of HIV-1-specific CD8+ cytotoxic T lymphocytes (CTL). Following antigen-specific activation, these mediators are secreted together, facilitating both lysis of virion-producing cells and the inhibition of free virus. In addition, RANTES, MIP-1alpha and MIP-1beta are secreted by CTL as a macromolecular complex containing sulphated proteoglycans. This association appears to have a functional significance, because heparan sulphate facilitates RANTES inhibition of HIV-1 infection of monocytes.

Cell Line, Transformed↗
PubMed1997

Activation of caspase-2 in apoptosis.

Members of the CED-3/interleukin-1beta-converting enzyme (ICE) protease (caspase) family are synthesized as proforms, which are proteolytically cleaved and activated during apoptosis. We report here that caspase-2 (ICH-1/NEDD-2), a member of the ICE family, is activated during apoptosis by another ICE member, a caspase-3 (CPP32)-like protease(s). When cells are induced to undergo apoptosis, endogenous caspase-2 is first cleaved into three fragments of 32-33 kDa and 14 kDa, which are then further processed into 18- and 12-kDa active subunits. Up to 50 microM N-acetyl-Asp-Glu-Val-Asp-aldehyde (DEVD-CHO), a caspase-3-preferred peptide inhibitor, inhibits caspase-2 activation and DNA fragmentation in vivo, but does not prevent loss of mitochondrial function, while higher concentrations of DEVD-CHO (>50 microM) inhibit both. In comparison, although the activity of caspase-3 is very sensitive to the inhibition of DEVD-CHO (<50 nM), inhibition of caspase-3 activation as marked by processing of the proform requires more than 100 microM DEVD-CHO. Our results suggest that the first cleavage of caspase-2 is accomplished by a caspase-3-like activity, and other ICE-like proteases less sensitive to DEVD-CHO may be responsible for activation of caspase-3 and loss of mitochondrial function.

Apoptosis↗
PubMed1997

Nosocomial infections in pediatric patients with burns.

BACKGROUND: Nosocomial infections (NI) are believed to occur more commonly in patients with burns than in patients undergoing surgery, but benchmark rates have not been well described, and widely accepted definitions of NI in patients with burns are not available. We present a clinically useful set of definitions for NI for the pediatric burn population and provide benchmark infection rates for NI at selected sites. METHODS: Centers for Disease Control and Prevention definitions were modified to more accurately describe nosocomial burn infection and secondary bloodstream infections (BSI) in the burn population. A surveillance system was developed and included calculation of NI rates by 1000 patient or device days, stratified into one of three risk groups (< 30% burn injury, 30% to 60% burn injury, and > 60% burn injury). All patients with acute burns admitted from January 1990 to December 1991 were included, and NI rates were calculated for burn infection, primary and secondary BSI, ventilator-related pneumonia and urinary catheter-related urinary tract infection (UTI). RESULTS: Overall 12.5% of patients with central venous catheters had development of primary BSI for a rate of 4.9/1000 central venous catheter-days. Incidence of secondary BSI was 5.8% of patients for a rate of 5.3/1000 patient-days. Incidence of burn infection was 10.1% of patients for a rate of 5.6/1000 patient-days. Incidence of ventilator-related pneumonia was 17.5% of patients for a rate of 11.4/1000 ventilator-days. Incidence of urinary catheter-related UTI was 17.9% of patients, for a rate of 13.2/1000 urinary catheter-days. When rates were stratified by risk groups, incidence increased with increasing burn size for secondary BSI (p < or = 0.0001) and urinary catheter-related UTI (p = 0.08), although rates based on number of patient-days or device-days more accurately reflected risk of infection over time. CONCLUSIONS: Infection remains a cause of significant morbidity and death for patients with burns. The definitions and benchmark rates reported here may be useful in evaluation of NI surveillance strategies and calculation of infection rates, which could then be used to evaluate current treatment modalities and improve outcomes for the burn population.

Burns↗
PubMed1996

Haptoglobin phenotyping by newly developed monoclonal antibodies. Demonstration of haptoglobin uptake into peripheral blood neutrophils and monocytes.

We have generated two IgG murine mAbs that recognize native human haptoglobin (Hp). These mAbs, 3A8 and 4B2, efficiently bind to the Hp complex regardless of the serum donor's phenotype. The specificity of mAb 3A8 was confirmed by immunoaffinity purification of 3A8-binding material from human serum and subsequent N-terminal amino acid sequencing of the invariant 40-kDa chain. mAb 3A8 and 4B2 were also reactive with cell-associated Hp when studied by immunocytochemistry. When human peripheral blood leukocytes were tested, 90% of the granulocytes and a lesser (and variable) fraction of monocytes displayed an intense intracytoplasmatic granular staining. This was confirmed by flow cytometric analysis of permeabilized leukocytes and by demonstrating the presence of Hp (of the expected Hp serum phenotype) in extracts of washed granulocytes by immunoblotting. Leukocytes obtained from a Hp phenotype 2-2 donor, incubated in culture medium supplemented with 10% serum from a donor possessing the Hp 1-1 phenotype, contained both Hp phenotypes when analyzed by immunoblotting after a 6-h incubation period. In addition, Hp was actively exocytosed by granulocytes following their exposure to Candida albicans. These observations suggest that exogenous Hp is concentrated within granulocytes and not synthesized de novo and is, in turn, exocytosed following neutrophil activation. Northern blotting analysis is consistent with the lack of haptoglobin gene transcription in granulocytes. These findings together with the earlier observations that Hp modulates granulocyte activity suggest that Hp levels may be enhanced locally at sites of inflammation to modulate granulocyte activity.

Amino Acid Sequence↗
PubMed1995

Inhibition of cytotoxic T-lymphocyte-triggered apoptosis by target cell surface-coupled aprotinin.

A variety of recent investigations have implicated granzymes A and/or B in the target cell nuclear injury which accompanies cytotoxic T-lymphocyte-mediated cytolysis. Since soluble antiproteases have had limited efficacy in inhibiting CTL-mediated lysis, we developed a method to couple aprotinin, a peptide inhibitor of serine proteases, to the surface of target cells. Aprotinin modified by N-succinimidyl 3-(2-pyridyldithio)propionate retained trypsin-inhibitory activity, and target cells modified with aprotinin had demonstrable cell surface trypsin-inhibitory activity. Flow cytometry demonstrated that aprotinin was detectable on the target cell surface but underwent modulation at a rather rapid rate. When radiolabeled, aprotinin-coupled target cells were studied in 1-2 hr CTL assays, 51Cr release was little affected, but 125IUdR release was reduced up to 75% compared to controls. Corresponding apoptosis analysed by agarose gel electrophoresis and direct cytologic visualization was similarly reduced. Thus, aprotinin bound to the surface of target cells selectively protected target cells against CTL-mediated nuclear injury, and may serve as a model for the development of novel inhibitors of CTL-mediated lysis.

Animals↗
PubMed1994

Neonatal serologic screening and early treatment for congenital Toxoplasma gondii infection. The New England Regional Toxoplasma Working Group.

BACKGROUND: Most infants with congenital Toxoplasma gondii infection have no symptoms at birth, but many will have retinal disease or neurologic abnormalities later in life. Early detection and treatment of congenital toxoplasmosis may reduce these sequelae. METHODS: In Massachusetts since January 1986, and in New Hampshire since July 1988, newborns have been screened for intrauterine infection with T. gondii by means of an IgM capture immunoassay of blood specimens routinely collected for screening for metabolic disorders. Congenital infection is confirmed by assays for specific IgG and IgM antibodies in serum from infants and their mothers. For this study, infants with serologic evidence of infection underwent extensive clinical evaluation and received one year of treatment. RESULTS: Through June 1992, 100 of 635,000 infants tested had positive screening tests. Congenital infection was confirmed in 52 infants, 50 of whom were identified only through neonatal screening and not through initial clinical examination. However, after the serologic results became available, more detailed examinations revealed abnormalities of either the central nervous system or the retina in 19 of 48 infants evaluated (40 percent). After treatment, only 1 of 46 children had a neurologic deficit (hemiplegia attributable to a cerebral lesion present at birth). Thirty-nine treated children had follow-up ophthalmologic examinations when one to six years old; four (10 percent) had eye lesions that may have developed postnatally (a macular lesion in one child and minor retinal scars in three). CONCLUSIONS: Routine neonatal screening for toxoplasmosis identifies congenital infections that are subclinical, and early treatment may reduce the severe long-term sequelae.

Antibodies, Protozoan↗
PubMed1994

Control of methicillin-resistant Staphylococcus aureus in a pediatric burn unit.

BACKGROUND: Control of methicillin-resistant Staphylococcus aureus (MRSA) is particularly difficult in burn units, which are often cited as sources of hospital-wide MRSA outbreaks. We developed a successful MRSA control program and document here its apparent effectiveness in controlling MRSA transmission in a pediatric burn unit. METHODS: An MRSA control program that included surveillance culturing, clinician feedback, flexible, site-specific isolation, and a list of known carriers was consistently applied in a pediatric burn unit through a 7-year period. Microbiology reports of MRSA isolates from patients and environmental surfaces and records of all patients from whom MRSA was isolated were reviewed. RESULTS: During calendar years 1985 through 1991, a total of 991 acutely burned children were admitted to the Boston unit of the Shriners Burns Institute. Forty MRSA cases (4%) were identified. One patient both had MRSA at admission and met our criteria for nosocomial MRSA. Of the remaining 39 patients, 11 had MRSA at admission and 28 had nosocomial MRSA. There were 17 wound infections, two cases of pneumonia, and two bloodstream infections. No deaths were attributed to MRSA sepsis. CONCLUSION: An MRSA control program including surveillance culturing, clinician feedback, flexible, site-specific isolation, and a list of known carriers is associated with a low rate of nosocomial MRSA in a pediatric burn unit.

Adolescent↗
PubMed1993

Assessment of ability of murine and human anti-lipid A monoclonal antibodies to bind and neutralize lipopolysaccharide.

The use of monoclonal antibodies (mAbs) directed to lipid A for the therapy of gram-negative sepsis is controversial. In an attempt to understand their biologic basis of action, we used a fluid-phase radioimmunoassay to measure binding between bacterial lipopolysaccharide (LPS) and two IgM mAbs directed to lipid A that are being evaluated for the treatment of gram-negative bacterial sepsis. Both antibodies bound 3H-LPS prepared from multiple strains of gram-negative bacteria when large excesses of antibody were used, although binding was modest and only slightly greater than control preparations. We also studied the ability of each anti-lipid A antibody to neutralize some of the biological effects of LPS in vitro. Despite large molar excesses, neither antibody neutralized LPS as assessed by the limulus lysate test, by a mitogenic assay for murine splenocytes, or by the production of cytokines interleukin (IL)-1, IL-6, or tumor necrosis factor from human monocytes in culture medium or in whole blood. Our experiments do not support the hypothesis that either of these anti-lipid A mAbs function by neutralizing the toxic effects of LPS.

Adult↗
PubMed1993

Flavobacterial sepsis in massively burned pediatric patients.

An experience with wound sepsis due to Flavobacterium meningosepticum in two pediatric burn patients is described. This organism, which is typically found in water and soil, generally has low pathogenicity but may become clinically important in immunocompromised hosts. It is typically resistant to antibiotics prescribed for infections caused by aerobic, gram-negative bacteria. When flavobacteria are suspected as pathogens, initial therapy should begin with ciprofloxacin, trimethoprim-sulfamethoxazole, or clindamycin.

Adolescent↗
PubMed1992

Differences between ATP-mediated cytotoxicity and cell-mediated cytotoxicity.

There has been considerable recent interest in the possible role of extracellular ATP in cell-mediated cytotoxicity. To investigate the similarities and differences between ATP-mediated lysis and CTL-mediated lysis, we studied in detail the ATP-mediated lysis of EL-4 cells, which are highly susceptible to lysis by extracellular ATP. ATP-mediated lysis was time and concentration dependent; most lysis occurred within 4 to 6 h of incubation. The kinetics of ATP- and cell-mediated lysis were similar. However, in contrast to CTL-mediated lysis, ATP-mediated lysis of EL-4 cells did not appear to be accompanied by characteristic chromosomal degradation (apoptosis). In order to compare these cytotoxic processes in greater detail, ATP-resistant clones were derived by growing EL-4 cells continuously in the presence of high concentrations of ATP. These cloned EL-4 lines showed marked resistance to ATP-mediated cytotoxicity across a wide range of concentrations but were as susceptible as the parent EL-4 cells to cell-mediated cytotoxicity by an alloreactive effector population from a MLC. Thus, there appear to be a number of differences between ATP-mediated and cell-mediated cytotoxicity in this system; most notable is the identification of cell lines that are resistant to ATP but susceptible to CTL-mediated lysis.

Adenosine Triphosphate↗