Major histocompatibility complex class I molecule expression is normal on peripheral blood lymphocytes from patients with insulin-dependent diabetes mellitus.

Recent work from one laboratory has shown, in both nonobese diabetic mice and humans, an association between insulin-dependent diabetes mellitus (IDDM) and quantitative difference in MHC class I molecule expression. This reported decrease in MHC class I molecule expression is very controversial in the nonobese diabetic mouse model of IDDM, but to our knowledge, it has not been evaluated by another group in human IDDM. To evaluate this question, we studied 30 patients with IDDM and 30 age- and sex-matched normal controls. MHC class I molecule expression was measured by flow cytometry with conformational-dependent MHC class I mAbs. The mean antigen density of MHC class I molecule expression in IDDM vs. normal control is 454+/-34 vs. 440+/-28 for lymphocytes and 1,440+/-117 vs. 1,494+/- 117 for monocytes, both P > 0.05. Three conformational-dependent MHC class I antibodies showed consistent results. To estimate the biological variation of MHC class I molecule expression in normal controls, we also studied 10 age- and sex-matched normal control pairs. Using X +/-SD of the percentage difference of mean antigen density in the normal control pairs as our definition of normal, we found that 70% (21/30) of IDDM patients had normal, 13% (4/30) of IDDM patients had decreased, and 17% (5/30) of IDDM patients had increased MHC class I molecule expression on lymphocytes. All IDDM patients showed normal MHC class I expression on monocytes. In conclusion, we find that there is no consistent decrease in MHC class I molecule expression on either lymphocytes or monocytes from patients with IDDM. The MHC class I molecule expression observed in IDDM patients is largely within the expected biological variation of MHC class I molecule expression that has been observed in normal controls.

Abnormalities in intramyocardial arteries detected in cardiac transplant biopsy specimens and lack of correlation with abnormal intracoronary ultrasound or endothelial dysfunction in large epicardial coronary arteries.

OBJECTIVES: We sought to determine whether abnormalities in small intramyocardial vessels could be detected on routine cardiac transplant biopsy specimens and whether these features correlate with intimal thickening by intracoronary ultrasound and endothelial dysfunction in large epicardial vessels. BACKGROUND: Variability in clinical presentation of allograft vasculopathy suggests differential involvement of large and small vessels. Intracoronary ultrasound and endothelial function studies detect large-vessel abnormalities but may not reflect changes in small intramyocardial arteries. The latter could be detected in routine cardiac biopsy specimens by histologic and immunohistochemical studies. METHODS: Thirty-nine cardiac transplant recipients underwent intracoronary ultrasound and acetylcholine studies 5 to 7 days after endomyocardial biopsy. Biopsy tissue was evaluated for coronary artery endothelial plumping and intimal thickening and increased immunostaining for fibronectin, tumor necrosis factor-alpha and receptor for hyaluronan-mediated motility. Large-vessel disease was assessed by calculating an average intimal index from intracoronary ultrasound of the left anterior descending coronary artery. Endothelial function was determined by quantitative coronary analysis after acetylcholine challenge. RESULTS: Coronary arteries were found in the biopsy tissue of 30 (76%) of the 39 patients who formed the study group. Fourteen of 30 patients had abnormal histologic findings. Immunohistochemical analysis for fibronectin, possible in 20 of 30 patients, was positive in 14 (70%) of 20 and correlated with abnormal histologic findings (p = 0.01). Immunostaining was positive for tumor necrosis factor-alpha and receptor for hyaluronan-mediated motility in 12 (40%) and 13 (43%) of 30 patients, respectively. All patients had intimal thickening by intracoronary ultrasound, but intimal index did not correlate significantly with small-artery disease by histologic or immunohistochemical analysis. Large-vessel endothelial dysfunction in 13 patients (43%) did not correlate with either abnormal ultrasound findings or small-vessel disease. CONCLUSIONS: Intramyocardial arteries are readily observed in biopsy specimens from cardiac transplant recipients and provide useful information about allograft vasculopathy. Lack of correlation between intramyocardial and epicardial vessel disease suggests discordant progression of allograft vasculopathy.

Increasing erythromycin resistance among group A streptococci causing tonsillitis in a tertiary care hospital in southern India.

Group A streptococci (GAS) (n = 1313) isolated from patients with clinical symptoms of pharyngitis or tonsillitis attending a tertiary care hospital in southern India during 1986-2002 were tested for susceptibility to penicillin and erythromycin. The overall erythromycin resistance rate was 2.7% (n = 36). During 1986-1993, erythromycin resistance was observed in only one (2%) isolate in 1987, but reappeared in 1994 (2.7%), increased to 5.8% in 1999, and reached a maximum frequency of 13.8% in 2002. All isolates were susceptible to penicillin. The data indicate the need for continued surveillance of susceptibility patterns among GAS isolates in order to monitor the development of antibiotic resistance.

Extending the B7 (CD80) gene family.

B7-1 and B7-2 are members of the immunoglobulin superfamily (IgSF) and important regulators of T cell-mediated immune responses. Despite sharing only limited sequence identity, B7-1 and B7-2 bind common receptors, CD28 and CTLA-4, on T cells and have similar functional properties. We have found that the extracellular V (ariable)-like domains of B7-1 and B7-2 share significant sequence similarities with 3 major histocompatibility complex (MHC)-encoded members of the IgSF: butyrophilin, myelin/oligodendrocyte glycoprotein, and the chicken MHC molecule, B-G. This raises the question whether there is an evolutionary link between the MHC, which encodes molecules regulating the antigen specificity of T lymphocyte responses, and B7 molecules, which co-stimulate these responses in antigen-nonspecific fashion.

Oral sumatriptan-associated transmural myocardial infarction.

A 43-year-old healthy man developed transmural myocardial infarction shortly after ingesting sumatriptan succinate 100 mg for migraine. Coronary arteriography revealed only minor irregularities in the left anterior descending artery. Oral sumatriptan should be used with caution in any patient with vascular risk factors and avoided in those with coronary artery disease or vasospasm.

CTLA4Ig treatment ameliorates the lethality of murine graft-versus-host disease across major histocompatibility complex barriers.

Graft-versus-host disease (GVHD), a pathological condition associated with BMT, results from activation of donor T lymphocytes by host tissues. CD28 and CTLA-4 are structurally related T cell receptors for members of the B7 (CD80) gene family, which transmit important costimulatory signals for T cell activation in vitro and in vivo. Here we have investigated the effects of CTLA4Ig, a soluble form of CTLA-4, on lethal GVHD in a murine model. Lethal GVHD was induced by transfer of parent C57BL/6 bone marrow and spleen cells into lethally irradiated (C57BL/6 x DBA/2)F1 recipients. Short courses of treatment with CTLA4Ig did not block engraftment, but prolonged survival of BMT recipients even when administration was delayed for 6 days after transplantation. CTLA4Ig-treated survivors of GVHD maintained body weight and did not exhibit visible signs of GVHD. However, treatment regimens that maximally prolonged survival did not detectably prevent T cell-mediated hematological abnormalities associated with GVHD, including pancytopenia and abnormal cellular composition of the spleen. Our data thus show that the lethality of acute GVHD in this model system is more dependent upon CD28/CTLA-4 costimulation than are other GVHD-associated abnormalities, and can be blocked for an extended period by brief treatment with CTLA4Ig.

The 'monocytic' cell line I937 displays typical B cell characteristics.

The monocytic cell line I937 was derived from U937 by sorting for cells with high expression of MHC class II molecules. Further analysis of these class II molecules revealed the presence of the HLA-DR3 subtype suggesting that the cell line was a potential candidate for testing antigen presentation to T cells restricted by HLA-DR3. We found that the T cell clones CFTS4:2.80 and CFTS4:2.6 with the required restriction element responded to the house dust mite antigen DPT presented by I937 but not U937, whereas CFTS4:3.1, which is not HLA-DR restricted, did not respond to either cell line. Subsequent analysis of surface markers on I937, however, indicated that the cell line is of B cell origin. In contrast to the parental cell line U937, I937 was tested negative for CD4, CD31 and CD64 but expressed CD19, CD21 and CD40. Although neither surface nor cytoplasmic Ig molecules were detected in either I937 or U937, Southern blot analysis revealed IgH gene rearrangement in I937. In addition, a fragment specific for Epstein-Barr virus nuclear antigen (EBNA2) was amplified in I937 by PCR technique. Therefore, we conclude that I937 is an EBV-transformed B cell line, presumably derived from the same donor and not as reported originally as a subline of U937, which expresses high MHC class II levels.

Improved specificity of myocardial thallium-201 single-photon emission computed tomography in patients with left bundle branch block by dipyridamole.

Reduced septal uptake of thallium-201 during exercise is frequently observed in patients with left bundle branch block (LBBB) and normal coronary arteries. This may reflect normal coronary autoregulation in response to lower septal oxygen demand; thus, dipyridamole, which uniformly exploits flow reserve, would be more accurate for diagnosis of coronary artery disease (CAD). Sixteen patients with LBBB underwent exercise and dipyridamole thallium-201 single-photon emission computed tomography and coronary angiography within 3 months. Sensitivity for detection of left anterior descending CAD (greater than 50% stenosis) was 0.83 for exercise and 1.00 for dipyridamole. Specificity was 0.30 (visual) or 0.20 (quantitative analysis) for exercise and 0.80 (visual) or 0.90 (quantitative) for dipyridamole (p less than 0.05). Dipyridamole combined with quantitative analysis also improved specificity of CAD detection overall (p less than 0.01). These data demonstrate that pharmacologic vasodilation is more accurate than exercise when diagnosing CAD by myocardial perfusion scintigraphy in patients with LBBB.

Myocardial infarction determined by technetium-99m pyrophosphate single-photon tomography complicating elective coronary artery bypass grafting for angina pectoris.

The incidence of acute myocardial infarction (AMI) complicating coronary artery bypass grafting (CABG) has previously been based on concordance of electrocardiographic, enzymatic and scintigraphic criteria. Technetium-99m pyrophosphate (Tc-PPi) single-photon emission computed tomography now enables detection of AMI with high sensitivity and specificity. Using this technique, perioperative AMI was detected in 12 of 58 patients (21%) undergoing successful elective CABG for stable angina pectoris. Stepwise multivariate logistic regression analysis was performed to compare the predictive value of preoperative (New York Heart Association class, left ventricular ejection fraction and use of beta blockers) and intraoperative (number of grafts constructed, use of internal mammary anastomoses, use of sequential saphenous vein grafts, smallest grafted distal vessel lumen caliber and aortic cross-clamp time) variables. Preoperative New York Association class (p = 0.04) and smallest grafted distal vessel lumen caliber (p = 0.03) were significant multivariate predictors of perioperative AMI. Only 1 perioperative patient with AMI (and 1 pyrophosphate-negative patient) developed new Q waves. Serum creatine kinase-MB was higher in patients with AMI by repeated measures analysis of variance (p = 0.0003). Five AMIs occurred in myocardial segments revascularized using sequential saphenous vein grafts, and 7 in segments perfused by significantly stenosed epicardial vessels with distal lumen diameter and perfusion territory considered too small to warrant CABG. At 6-month follow-up, the mean left ventricular ejection fraction increased from 0.61 to 0.65 in Tc-PPI-negative patients (p = 0.01), but not in perioperative patients with AMI.(ABSTRACT TRUNCATED AT 250 WORDS)

Signal transduction in lymphocyte activation through crosslinking of HLA class I molecules.

The inhibitory effect of anti-HLA class I monoclonal antibodies on lymphocyte proliferation has been well documented. However, recent data suggest that anti-HLA class I monoclonal antibodies can enhance lymphocyte proliferation via both anti-CD3-induced (1,2) and anti-CD2-induced (3) activation pathways. Here we demonstrate that both inhibition and activation can be regulated by the degree of aggregation of HLA class I antigens. Crosslinking of monoclonal antibodies specific for HLA-A, HLA-B, or monomorphic determinants (using anti-IgG2 and/or anti-Ig kappa "second step" monoclonal antibodies) increased the capacity of anti-HLA class I monoclonal antibodies to inhibit phytohemagglutinin-induced proliferation. However, the cytosolic free calcium concentration was increased in CD4+ cells, CD8+ cells, B cells, and CD16+ cells when anti-HLA class I monoclonal antibodies were crosslinked, suggesting that an activation signal was generated by aggregation of the corresponding antigens. Indeed, inositol 1,4,5-trisphosphate could be detected in peripheral blood lymphocytes following crosslinking of anti-HLA class I monoclonal antibodies. Class I aggregation also induced proliferation of peripheral blood mononuclear cells in the presence of submitogenic doses of phorbol 12-myristate 13-acetate. Strong conditions of crosslinking (monomorphic monoclonal antibody plus both anti-IgG2 and anti-Ig kappa) induced CD25 expression and responsiveness to recombinant interleukin 2. Our results suggest that aggregation of HLA class I antigens primed cells to become activated in the presence of progression signals including phorbol 12-myristate 13-acetate, recombinant interleukin 2, or anti-CD5 plus anti-CD28 monoclonal antibodies.

Myocardial vacuolization, a marker of ischemic injury, in surveillance cardiac biopsies posttransplant: Correlations with morphologic vascular disease and endothelial dysfunction.

Allograft vasculopathy (AV) causes intimal thickening with progressive luminal obstruction, endothelial dysfunction, and abnormal vasomotion. Subendocardial vacuolization indicating ongoing ischemia was observed at autopsy in transplanted hearts with severe AV. Whether myocyte vacuolization can be observed with lesser degrees of AV in cardia transplant patients has not been reported. Thirty-nine cardiac transplant patients without flow-limiting disease in large epicardial arteries underwent invasive assessment of AV. Eight to 10 segments of the left anterior descending artery were analyzed by intracoronary ultrasound, and an average intimal index was calculated. Endothelial response to acetylcholine was assessed with serial quantitative angiography. Endomyocardial biopsies taken 5 to 7 days prior to the invasive studies were histopathologically reviewed for the presence of small intramyocardial arteries and myocyte vacuolization. Myocyte vacuolization was evident in biopsies from 20 patients (51%). Intramyocardial arteries were observed in 30 cases (76%); 14 had abnormal arteries. All patients had some degree of intimal thickening by intracoronary ultrasound, and 7 (17 %) had severely abnormal average intimal index (>0.2). Endothelial dysfunction was present in 23 patients (58%). Vacuolization failed to show an association with abnormal small artery histology or large epicardial artery ultrasound disease. However, a significant association between vacuolization and endothelial dysfunction was observed (p = 0.05). Myocyte vacuolization, possibly indicating ischemic injury, is common in biopsies from cardiac transplant patients and is associated with abnormal acetylcholine response in large epicardial arteries. We speculate that myocyte vacuolization may be caused at least in part by impaired coronary flow associated with endothelial dysfunction.

Invasive properties of south Indian strains of Streptococcus pyogenes in a HEp-2 cell model.

The objective of this study was to consider the invasive properties of Streptococcus pyogenes in human pharyngeal epithelial cells, and to correlate these with their clinical significance. Clinical isolates of S. pyogenes obtained from blood cultures over a period of 10 years, and throat and skin isolates from a community-based study, were used in this investigation. The S. pyogenes isolates were inoculated in HEp-2 cells and subsequently treated with antibiotics to kill the extracellular bacteria. The cells were then lyzed, and a colony count was carried out to check for invasion. The throat and skin isolates had 45.7%, 25.7% and 28.5% of low, intermediate and high invasion efficiencies, respectively, while 80%, 8.6% and 11.4% of the blood isolates had low, intermediate and high invasion efficiencies. We concluded that the throat and the skin isolates from superficial infections were more invasive than the blood isolates, which is an interesting and paradoxical feature.

Microbiological diagnosis of streptococcal pharyngitis: lacunae and their implications.

Post-streptococcal sequelae, especially acute rheumatic fever/rheumatic heart disease continue to occur in significant proportions in many parts of the world. Despite several attempts with various intervention strategies, little success has been achieved in the control of acute rheumatic fever/rheumatic heart disease in India. The success of the control programmes depends upon timely primary prophylaxis with benzathine penicillin for which a microbiological confirmation of group A streptococcal pharyngitis is essential. Isolation of beta hemolytic streptococci from throat cultures and their identification as GAS in the laboratory, clinches the microbiological diagnosis while demonstration of a 'significant rise' in antibody titers such as Anti-streptolysin O and Anti-deoxyribonuclease B differentiates it from a group A streptococcal carrier state or pharyngitis of a viral etiology. Despite the easiness with which these can be achieved, many laboratories in India are not equipped to do so. Enhancing bacteriological and serological facilities in laboratories across the country will drastically improve the clinician's ability to diagnose bona fide GAS pharyngitis and help to institute penicillin prophylaxis at the appropriate time. This will go a long way in enhancing the compliance to penicillin prophylaxis which is the cornerstone of any RF/RHD control program.

Non tuberculous mycobacteria isolated from clinical specimens at a tertiary care hospital in South India.

PURPOSE: This is a retrospective analysis of the isolation rates of nontuberculous mycobacteria (NTM) from various clinical specimens and their antimicrobial susceptibility patterns. METHODS: All NTM isolated between 1999 and 2004 at Christian Medical College, Vellore, South India, were identified with various biochemical tests. Antimicrobial susceptibility test for all NTM was performed by standard methods. RESULTS: A total of 32,084 specimens were received for culture, of which 4473 (13.9%) grew acid fast bacilli (AFB). Four thousand three hundred (96.1%) of the AFB were M. tuberculosis while 173 (3.9%) were NTM. Of the 173 NTM, 115 (66.5%) were identified to the species level. Pus, biopsy specimens and sputum specimens yielded most of the NTM of which M. chelonae (46%) and M. fortuitum (41%) accounted for majority of them. M. chelonae and M. fortuitum, showed highest susceptibility to amikacin (99.2%). NTM were repeatedly isolated from seven sputum specimens, 15 biopsy and pus specimens, two CSF and two blood cultures. Six were isolated from patients with AIDS and five from post transplant patients. CONCLUSIONS: The isolation of NTM from various clinical specimens is reported in this study to highlight the associated diseases and therapeutic options in these infections.

Incidence of carbapenem resistant nonfermenting gram negative bacilli from patients with respiratory infections in the intensive care units.

Resistance to carbapenems is commonly seen in nonfermenting gram negative bacilli (NFGNB). We document herein the prevalence of carbapenem resistance in NFGNB isolated from patients with respiratory tract infections in the intensive care units (ICUs). A total of 460 NFGNB were isolated from 606 endotracheal aspirate specimens during January through December 2003, of which 56 (12.2%) were found to be resistant to imipenem and meropenem. Of these, 24 (42.8%) were Pseudomonas aeruginosa , 8 (14.2%) were Acinetobacter spp. and 24 (42.8%) were other NFGNB. Stringent protocols such as antibiotic policies and resistance surveillance programs are mandatory to curb these bacteria in ICU settings