Frequency of Panel Reactive Antibodies (PRA) among Renal Transplant Recipients and its Effect Modifiers.

OBJECTIVE: To determine frequency of panel reactive antibodies among renal transplant recipients and its effect modifiers. STUDY DESIGN:   A cross-sectional study. PLACE AND DURATION OF STUDY: Department of Immunology, Armed Forces Institute of Pathology from October 2016 to October 2017. METHODOLOGY: One hundred and sixty-two (162) patients, who were referred to Department of Immunology for pre-transplant workup for kidney transplantation of both genders and Pakistani nationality. Informed consents were taken and detailed history were recorded. Frequency and percentages were calculated for panel reactive antibodies, blood transfusion, pregnancy and previous transplant were noted and Chi-square test was applied. RESULTS: One hundred and sixty-two (162) patients including 141 males and 21 females were analysed and 48 patients (30%) were positive for panel reactive antibodies (PRA). Of 141 male patients analyzed, 35 were positive for PRA, which were about 25%. Twenty-one females were tested for PRA and 13 female patients were positive that is about 62% of the analysed population. Out of the total 141 males, 20 (14%) had blood transfusion and of these 11 (55%) were positive for PRA. Without history of transfusion, only 9 (7%) were positive for PRA. Out of 21 females, 10 were positive for blood transfusion, out of which 6 (60%) were positive for PRA. Without history of blood transfusion, 7 (64%) were positive for PRA. Out of 21 females, 20 had history of pregnancy. Out of whom, 13 (65%) were positive for PRA. Two patients (one male and one female) were with history of previous transplant and both were positive for PRA. CONCLUSION: A significant number of patients were sensitised with panel reactive antibodies waiting for renal transplant. The PRA was more common in recipients who were prone to effect modifiers such as pregnancy, blood transfusion and re-transplant. These risk factors were mostly present in combination, which also suggests their synergistic effects on PRA synthesis. Key Words: Blood transfusion, Effect modifiers, Panel reactive antibodies, Pregnancy, Kidney transplant, Re-transplant.

Distribution of HLA-B*27 Subtypes in Patients with Ankylosing Spondylitis in Local Population.

OBJECTIVE: To compare the distribution of HLA-B*27 subtypes in healthy controls and in ankylosing spondylitis (AS) patients of different ethnic groups from Pakistan. STUDY DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from April 2016 to October 2017. METHODOLOGY: Forty-nine HLA-B*27 positive, unrelated AS patients and 18 HLA-B*27 positive healthy BMT/renal transplant donors were selected for this study. Typing of the HLA-B27 alleles was performed by the polymerase chain reaction-sequence-specific primer (PCR-SSP). RESULTS: There was a wide number of HLA-B*27 subtypes and an elevated frequency of the B*2707 allele in the AS patients. The allele B*2706 seems to have a protective role in the population studied because it was found only in the healthy controls. HLA-B*27:03 and 07 were found predominant subtypes in Punjabis and Pathans, respectively. CONCLUSION: There were no significant differences for the distribution of B*27 subtypes between patients and controls (p >0.05).

Leukocyte Adhesion Deficiency Type 1 with Low Expression of CD 11b.

Leukocyte adhesion deficiency type 1 (LAD-1) is a rare autosomal recessive disorder caused by mutations in the gene that codes for CD18, the beta chain of beta-2 integrins, located on the long arm of chromosome 21. This defect results in failure of leukocyte migration to the site of infection due to the absence of surface integrins. Leukocyte adhesion deficiency should be suspected in any patient with recurrent infections, impaired wound healing, history of delayed umbilical cord separation, periodontitis, leukocytosis, recurrent soft tissue and oral infections. Diagnosis is based on the analysis of neutrophils for the surface expression of CD18, CD11a, CD11b and CD11c by flow cytometry. Here, we present a 55-day male infant with umbilical cord separation on the 10th day of life and no history of infection, who was identified with LAD-1 with low expression of CD11b. The purpose of performing LAD flow cytometric analysis in this patient was to screen him for LAD-1 as his elder brother had LAD-1 and one elder sister died undiagnosed with recurrent skin and chest infections at 8 months of age.

Significance of Testing Anti-Thyroid Autoantibodies in Patients with Deranged Thyroid Profile.

BACKGROUND: We hypothesized that anti-thyroid antibodies are more often positive in individuals with deranged thyroid profile. METHODS: This prospective cohort was done in Immunology Department, Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from Jan 2017 to Oct 2017. All the samples that were referred to us for testing anti-thyroid antibodies (anti-TPO or anti-TG antibodies) and thyroid profile were included in the study. There were no exclusion criteria. Tests for anti-thyroid antibodies were performed by ELISA and thyroid profile by chemiluminescence. SPSS 23.0 was used for statistical analysis. RESULTS: Over a course of a ten-month study period, we received a total of 316 serum samples for anti-TPO/TG antibodies along with thyroid profile testing (TSH). These included 115 males (36.4%) and 201 females (63.6%). Their age ranged from 3 to 89 years (mean ± SD, 42.22 ± 18.09). Anti-TPO antibodies were more often positive when TSH was deranged (p value 0.001). Anti-TPO antibodies are more often raised in females, in terms of both prevalence (p 0.001) and mean rank (p 0.002). CONCLUSION: As anti-thyroid antibodies are more often present when TSH is deranged, such individuals should be screened for anti-thyroid antibodies. This importance of screening is compounded by the fact that anti-thyroid antibodies may be positive in a significant percentage of elderly people.

Determination of Specificity and Pattern of Antinuclear Antibodies (ANA) in Systemic Rheumatic Disease Patients Positive for ANA Testing.

OBJECTIVE: To determine probability of finding antinuclear antibodies (ANA) and anti extractable nuclear antigens (ENA) positive samples and associating ANA patterns with anti-ENA reactivities among a consecutive cohort of samples of systemic rheumatic disease patients referred for ANA testing. STUDY DESIGN: Prospective cohort study. PLACE AND DURATION OF STUDY: Immunology Department, Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from January to June 2016. METHODOLOGY: All the samples referred for ANA testing with clinical suspicion of systemic rheumatic disease were included. After screening, ANA positive samples were subjected to anti-ENA antibodies testing (including anti-SSA, antiSSB, anti-Sm, anti-RNP, anti-SCL-70 and anti-Jo-1 antibodies) and ANA pattern and titer determination. RESULTS: Of 4,347 samples received, 397 were positive for ANA (9%). Of 397, 96 (24%) samples positive on ENA screen were tested for anti-ENA reactivity. Anti-SSA antibodies were found in 59 samples. Commonest ANA patterns were coarse and fine speckled (43 and 22 samples of 81 tested), while majority of samples carried ANA in titers of 1:40 and 1:80 (22 and 18 samples of 81 tested). No specific ANA pattern was associated with any particular anti-ENA reactivity. CONCLUSION: Among samples/patients referred for investigations of autoimmune disorders, probability of finding positive ANA is approximately 9%. Of these 9%, about 24% also show reactivity against ENA. Commonest ANA pattern is coarse speckled and majority of such patients carry ANA in titers ranging from 1:40 to 1:80. Commonest ENA reactivity was against SSA.

Identification of HLA Class I Misreads/Dropouts Using Serological Typing, in Comparison with DNA-based Typing.

Serology and DNA techniques are employed for Human Leukocyte Antigen (HLA) typing in different transplant centers. Results may not always correlate well and may need retyping with different technique. All the patients (with aplastic anemia, thalassemia, and immunodeficiency) and their donors, requiring HLA typing for bone marrow transplant were enrolled in the study. Serological HLA typing was done by complement-dependent lymphocytotoxicity while DNA-based typing was done with sequence specific primers (SSP). Serology identified 167 HLA A and 165 HLA B antigens while SSP in same samples identified 181 HLA A and 184 HLA B alleles. A11 and B51 were the commonest antigens/alleles by both methods. There were a total of 21 misreads and 32 dropouts on serology, for both HLA A and B loci with HLA A32, B52 and B61 being the most ambiguous antigens. Inherent limitations of serological techniques warrant careful interpretation or use of DNA-based methods for resolution of ambiguous typing.

Severe combined immunodeficiency due to adenosine deaminase deficiency.

Severe Combined Immunodeficiency is the term applied to a group of rare genetic disorders characterised by defective or absent T and B cell functions. Patients usually present in first 6 months of life with respiratory/gastrointestinal tract infections and failure to thrive. Among the various types of severe combined immunodeficiency, enzyme deficiencies are relatively less common. We report the case of a 6 years old girl having severe combined immunodeficiency due to adenosine deaminase deficiency.

HLA DRß1 alleles in Pakistani patients with rheumatoid arthritis.

OBJECTIVE: To determine frequencies of HLA DRß1 alleles in rheumatoid arthritis in Pakistani patients. STUDY DESIGN: Cross sectional/analytical study. PLACE AND DURATION OF STUDY: Department of Immunology, Armed Forces Institute of Pathology, Rawalpindi in collaboration with Rheumatology departments of Military Hospital, Rawalpindi and Fauji Foundation Hospital, Rawalpindi, from January 2009 to January 2010. METHODOLOGY: HLA DRß1 genotyping of one hundred Pakistani patients, diagnosed as having RA as per American College of Rheumatology revised criteria 1987, was done. HLA DRß1 genotyping was carried out at allele group level (DRß1*01-DRß1*16) by sequence specific primers in RA patients. Comparison of HLA DRß1 allele frequencies between patients and control groups was made using Pearson's chi-square test to find possible association of HLA DRß1 alleles with RA in Pakistani rheumatoid patients. RESULTS: HLA DRß1*04 was expressed with significantly increased frequency in patients with rheumatoid arthritis (p <0.05). HLA DRß1*11 was expressed statistically significantly more in control group as compared to rheumatoid patients indicating a possible protective effect. There was no statistically significant difference observed in frequencies of HLA DRß1 allele *01, DRß1 allele *03, DRß1 allele *07, DRß1 allele *08, DRß1 allele *09, DRß1 allele*10, DRß1 allele *12, DRß1 allele *13, DRß1 allele *14, DRß1 allele *15 and DRß1 allele *16 between patients and control groups. CONCLUSION: The identification of susceptible HLA DRß1 alleles in Pakistani RA patients may help physicians to make early decisions regarding initiation of early intensive therapy with disease modifying anti rheumatic medicines and biological agents decreasing disability in RA patients.

Smear-negative pulmonary tuberculosis and lymphocyte subsets.

OBJECTIVE: To correlate and quantitate lymphocyte subsets with clinically diagnosed smear-negative pulmonary tuberculosis and severity of disease. DESIGN: Case-control study. PLACE AND DURATION OF STUDY: Military Hospital and Armed Forces Institute of Pathology Rawalpindi. 1999-2000. SUBJECTS AND METHODS: Freshly diagnosed, well-characterized smear-negative patients (n=15) of pulmonary tuberculosis were selected. Non-induced three-consecutive negative smears of sputum with simultaneous culture for AFB for 6-8 weeks, positive Mauntoux test (Z10 mm), blood complete picture with ESR and chest x-rays were done. Selected panel of monoclonal antibodies against specific CD markers were used. Statistical analysis done by student t-test or Mann-Whitney rank-sum test with the help of Sigma State software. RESULTS: Hemoglobin and total lymphocyte counts were significantly reduced whereas total leukocyte counts with absolute neutrophil counts were increased. Fraction of CD4+ and CD8+ T lymphocytes with HLA-DR expression was reduced while no significant change in rest of the TB and NK lymphocytes. CONCLUSION: Low hemoglobin level, high neutrophil count and low total lymphocytes suggest possible direct relationship with extent of disease. The number of activated CD4+, CD8+, ab and gd TCR T cells have tendency to increase during Mycobacterium infection. This seems to have a potential of being a good, non-invasive prognostic indicator for patients with pulmonary tuberculosis.