Anti NMDA receptor antibody encephalitis in Pakistan: Clinicopathological features and treatment outcomes.

Anti-NMDA receptor antibody encephalitis (anti-NMDAR Encephalitis) is the most common subtype of autoimmune encephalitis in which IgG antibodies directed against NR1 subunit of NMDA receptors are present. It is a potentially lethal encephalitis which responds favourably to timely immunosuppressive therapy. If untreated, its progression leads from delusions, paranoia, movement disorder, memory deficit and seizures into a state of unresponsiveness with autonomic instability and even death. We present clinicopathological features, treatment and outcomes of eight autoantibodyproven cases of anti-NMDAR Encephalitis. There were 7 females and 1 male with a mean age of 15 years (age range: 1 to 28 years). Clinical features included seizures, altered consciousness, memory deficit, delusions, paranoia and hallucinations. Hyperactivity and irritability were prominent features among the children. Patients treated with immunosuppressive therapy including steroids, IVIg, plasmapheresis and Rituximab, recovered completely within a month of therapy. Whereas patients who received only steroids as immunosuppressive therapy suffered from residual brain damage.

C1Q Nephropathy: A Multifaceted Disease With Infrequent Diagnosis.

BACKGROUND: C1q nephropathy (C1qN) is a rare glomerulopathy, with a very low prevalence world wide varying from 0.2 to 2.5%. Even though more than three decades have passed since this entity was first explained, still, it remains a dilemma for many due to the rarity of this lesion. This study was carried out principally to determine the clinical presentation, morphologic features and distribution of C1qN in our region based on renal biopsies studied by light microscopy (LM), and immunofluorescence (IF) so that this entity is better understood both by nephrologists and pathologists as no such study has ever been conducted in Pakistan to our knowledge. METHODS: It was a cross-sectional study carried out from 1st January 2012 to 30th December 2016 in Histopathology department, Shifa International Hospital. All cases diagnosed as C1q nephropathy were retrieved from the hospital's computerized database. Their clinical profiles, morphology and immunohistochemical profiles were studied.. RESULTS: Over this period a total of 31 cases were diagnosed with C1qN. Mean age of the patients was 32.09±18.66 years. The most common clinical presentation was nephrotic syndrome seen in 22 (71%) patients. The most frequent morphological pattern seen was minimal change disease (MCD) in 13 (41.9%) cases. All cases showed dominant 22 (71%) or codominant 9 (42.9%) mesangial±membranous C1q deposition. No correlation was found (p-value >0.05) between morphological pattern and clinical presentation of the disease or immunofluorescence findings. CONCLUSIONS: C1qN is a rare entity which is primarily diagnosed on the basis of immunofluorescence findings with a dominant or codominant fluorescent intensity for C1q. It is recommended that C1qN is sought for preferably with immunofluorescence staining of biopsies for immune reactants, especially for C1q. Studies from this part of the world are strongly recommended to predict clinical outcome and treatment options.

T cell reactivity to Collagen II as a possible prognostic marker in patients with rheumatoid arthritis.

OBJECTIVE: To compare the frequency and the functional state of the collagen II reactive T cells with disease activity in rheumatoid arthritis patients and healthy controls. METHODS: This case-control cross-sectional study was carried out at the Department of Immunology; Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from June to October 2014. Rheumatologist from Rehmat Noor Rheumatology Clinic, a private health facility of the city, was requested to send in patients with clinical diagnosis of rheumatoid arthritis. Samples were obtained and relevant investigations were carried out. Data were compared with a group of age and gender-matched healthy subjects. T cell proliferative response was assessed against bovine collagen II by measuring incorporation of bromodeoxyuridine into deoxyribonucleic acid of proliferating cells and by expression of CD25 on proliferating cells as percentage of CD3+/bromodeoxyuridine+ and CD3+/CD25+ T-cells, respectively. Among the patients, the frequency of T cells with disease activity was compared. Patients were classified into groups of mild, moderate and severe disease and frequency of CD3+/bromodeoxyuridine+, frequency of CD3+/CD25+ cells, mean fluorescent intensity of bromodeoxyuridine-fluorescein isothiocyanate and mean fluorescent intensity of CD25-fluorescein isothiocyanate were compared in the groups. RESULTS: Of the 60 subjects, 30(50%)were patients and 30(50%) were controls. Of the patients, 5(16.66%) were males and 25(83.33%) were females with an overall mean age of 42±12 years. The mean age of the controls was 41±9.28 years. Mean disease duration of the patients was 10.5 ± 4.2 years. Percentage of CD3+/CD25+ cells and CD3+/bromodeoxyuridine+ cells stimulated with collagen II, in patients was much higher than the controls(p<0.05).Statistically significant differences were observed when frequency of CD3+/bromodeoxyuridine+ cells and CD3+/CD25+ cells was compared among the mild, moderate and severe patient groups (p<0.05). CONCLUSIONS: Collagen II was found to be an important auto antigen in joints of rheumatoid arthritis patients.

Discordant disease expression of neonatal lupus erythematosus in twins.

Neonatal lupus erythematosus is an autoimmune disease resulting from the trans-placental passage of maternal anti-SSA/Ro, anti-SSB/La, and less frequently anti-RNP antibodies to the foetus. At the time of diagnosis 50% of mothers are asymptomatic. Neonatal manifestations of this multisystem disease may include congenital heart block, cutaneous lesions and haematological abnormalities. We present the case of congenital neonatal lupus erythematosus in non-identical twins, showing variability in clinical manifestation of this disease,despite receiving the same level of antibodies from the mother. This case adds to the growing body of evidence about the role of genetics and other feto-maternal contributing factors in addition to the presence of auto antibodies. It raises interesting questions about discordant disease expression in offspring's of the same mother.

Chronic granulomatous disease.

Chronic granulomatous disease is a rare inherited disorder characterised by inability of phagocytes to generate reactive oxygen species needed for intracellular killing of phagocytosed microorganisms. We report the case of an 8-month-old male child with recurrent chest infections and perianal abscess that had no response to conventional antibiotic treatment. His two elder brothers died due to similar complaints at the ages of 4 and 5 months. Four elder sisters were healthy and alive. This history indicated that the patient might have X-linked chronic granulomatous disease. A definite absence of superoxide activity in the patient's granulocytes detected by dihydrorhodamine test and nitroblue tetrazolium dye reduction test confirmed this diagnosis.

HLA- DR Alleles in Pakistani Patients of Pemphigus Vulgaris.

OBJECTIVE: To determine frequency of HLA-DR alleles in Pakistani patients of pemphigus vulgaris in comparison with local healthy controls. STUDY DESIGN: Cross-sectional, comparative study. PLACE AND DURATION OF STUDY: Department of Immunology, Armed Forces Institute of Pathology (AFIP), Rawalpindi, from January 2011 to January 2014. METHODOLOGY: Twenty eight patients with biopsy proven diagnosis of pemphigus vulgaris referred from Department of Dermatology, Military Hospital, Rawalpindi were included. Patients were compared with a group of 150 unrelated local healthy subjects. DNA was extracted from peripheral blood collected in Tri-potassium EDTA. HLA-DRB1 typing was carried out on allele level (DRB1*01--DRB1*16) using SSP (sequence specific primers). HLA type was determined by agarose gel electrophoresis and results recorded. Phenotype frequency of various alleles among patient group and control group was calculated by direct counting and significance of their association was determined by Fisher's exact test/ Chi square test. RESULTS: A total of 12 male and 16 female patients, with age ranging from 21 to 34 (mean 23.4 years) were genotyped for HLA-DRB1 loci. A statistically significant association of the disease with HLA-DRB1*04 was observed (50% versus 20.7% in controls, p < 0.05). CONCLUSION: There is a strong association of HLA-DRB1*04 with pemphigus vulgaris in Pakistani population.

Frequency of intrinsic factor antibody in megaloblastic anaemia.

OBJECTIVE: To evaluate the presence of intrinsic factor antibody in vitamin B12 deficient patients. STUDY DESIGN: Cross-sectional, observational study. PLACE AND DURATION OF STUDY: Fauji Foundation Hospital, Foundation University Medical College and Armed Forces Institute of Pathology, Rawalpindi, from January 2011 to June 2012. METHODOLOGY: A total of 120 patients of megaloblastic anaemia were selected on the basis of low serum vitamin B12 level. The intrinsic factor antibody tests were performed by ELISA method. The patients were considered positive or negative on the basis of presence or absence of intrinsic factor antibody respectively. The data was analyzed by using SPSS version 14. RESULTS: Pernicious anaemia with intrinsic factor deficiency was found in 13.3% in 120 vitamin B12 deficient patients. The mean age of patients of pernicious anaemia was 41.5 years, with a male to female ratio of 1:2.5. It was relatively more common in older age (17% in age more than 60 years) as compared to other age groups. CONCLUSION: Frequency of pernicious anaemia in megaloblastic anaemia was 13.3%. The male to female ratio was 1:2.5 and it was relatively more common in age group of more than 60 years.

HLA-DR alleles among Pakistani patients of coeliac disease.

OBJECTIVES: To investigate whether certain DR alleles might also contribute to the genetic susceptibility among Coeliac disease patients in Pakistan. METHODS: The case-control study was conducted at the Military Hospital, Rawalpindi, from October 2011 to January 2012, and analysed 25 children diagnosed to have coeliac disease as per the criteria set by the European Society of Paediatric Gastroenterology and Nutrition, which included histopathological alterations in duodenal biopsies, clinical response to gluten withdrawal, and presence of anti-endomyseal antibodies. Patients were compared with a group of 150 healthy subjects. Dioxyribonucleic acid was extracted from peripheral blood collected in ethylenediaminetetraacetic acid.K3. Human leukocyte antigen DRB1 typing was carried out on allele level (DRB1*01--DRB1*16) using sequence specific primers. Human leukocyte antigen type was determined by agarose gel electrophoresis and results were recorded. Phenotype frequency of various alleles among the patient group and the control group was calculated by direct counting, and significance of their association was determined by Fisher Exact Test. RESULTS: A total of 11 (44%) female paediatric coeliac patients in age range 1-9 (mean 7.2 +/- 4.8 years) and 14 (56%) male paediatric patients in the age range 6-14 (mean 8.6 +/- 5.1 years) were genotyped for HLA-DRB1 loci. A statistically significant positive association of the disease with HLA-DRB1*03 (n = 23; 92% versus n=31; 21% in controls, p < 0.01) was observed. CONCLUSION: HLA-DRB1*03 is associated with increased risk of developing coeliac disease.

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.

Clinical, histopathological and immunofluorescent findings of IgA nephropathy.

BACKGROUND: IgA nephropathy, a prevalent disease in Asia, is considered the main cause of end stage renal disease among primary glomerular disease. OBJECTIVE: To determine the frequency of different clinical, histopathological and immunofluorescent characteristics of IgA nephropathy. METHODS: Renal biopsies of 376 patients were received for immunofluorescent and for histopathological studies. Biopsies were stained with fluorescene isothyocyanate (FITC) labeled antibodies against IgG, IgA, IgM, C3, C4 and fibrinogen for fluorescent microscopy. For histopathological examination, the specimens were stained with hematoxylin and eosin, periodic acid schiff and methanamine silver stains for light microscopy. RESULTS: IgA nephropathy was diagnosed in 39 cases (10.4%) with a mean age 31.5 years and a male to female ratio of 2.8:1. The disease was observed in 11(29.7%) patients aged 21-30 years, followed by 8 patients (21.6%) aged 11-20 years group. Nephrotic range proteinuria was the most common laboratory finding which was detected in 11 patients (37%). Mesangioproliferative glomerulonephritis was the most common histopathological finding which was found in 7 patients (35%). IgA with other immunoglobulins and complements were deposited in 28 specimens (71.8%) as detected by immunofluorescence. CONCLUSION: IgA nephropathy is common in young people and one third of it results in end stage renal disease. We suggest that Immunofluorescent assay can be considered for the conclusive diagnosis of IgA nephropathy in young patients presenting with proteinuria/hematuria.

Human leukocyte antigen class II susceptibility conferring alleles among non-insulin dependent diabetes mellitus patients.

OBJECTIVE: To determine the frequency of Human Leukocyte Antigen (HLA) class II susceptibility conferring alleles among type 2 Diabetes mellitus patients, in comparison with healthy controls. STUDY DESIGN: Cross-sectional comparative study. PLACE AND DURATION OF STUDY: Department of Immunology, Armed Forces Institute of Pathology, Rawalpindi, from January 2009 to April 2010. METHODOLOGY: Patients with non-insulin dependent Diabetes mellitus meeting World Health Organization criteria were studied. These were compared with age and gender matched healthy control subjects. For each subject (patients as well as controls), DNA was extracted from ethylene diamine tetra-acetate sample and HLA class II DRB1 typing was carried out at allele group level (DRB1*01-DRB1*16) by sequence specific primers. Human leukocyte antigen DRB1 type was determined by agarose gel electrophoresis and results were recorded. Frequencies were determined as number of an allele divided by total number of alleles per group; p-value was computed using Pearson's chi-square test. RESULTS: Among the 100 patients, there were 63 males and 37 females with 68 controls. A total of 13 different HLA DRB1 alleles were detected, with DRB1*15 being the commonest in both the groups. The allele DRB1*13 had statistically significant higher frequency in patient group as compared to controls (p = 0.005). CONCLUSION: HLA DRB1*13 was found with a significantly increased frequency in non-insulin dependent Diabetes mellitus.

Leukocyte adhesion defect.

Leukocyte adhesion defect (LAD) is a rare, autosomal recessive primary immunodeficiency disorder of phagocytes, in which there is defective aggregation at the site of infection due to the absence of surface integrins. Diagnosis is based primarily on flowcytometric analysis of neutrophils for the surface expression of CD11, CD18 and CD15s. We describe here a case of a 7-months-old boy who presented with a characteristic history of recurrent infections, marked leukocytosis and delayed separation of umbilical cord. The diagnosis was established by demonstration of the absence of integrins on the surface of patient's neutrophils by flowcytometric analysis.

Chronic granulomatous disease.

Chronic granulomatous disease (CGD) is an X-linked/ autosomal recessive primary immunodeficiency disorder characterized by recurrent infections. The diagnosis is primarily based on simple Nitrobluctetrazolium dye reduction test. We describe here an unusual case of an 8 year old girl, as the disease is X-linked in most of the cases.

CD5-positive acute lymphoblastic leukemia.

CD5-positive B-ALL is a rare variant of Acute Lymphoblastic Leukemia (ALL). In literature, only three cases have been reported so far. This fourth case report describes a young lady who was diagnosed as ALL (L-2) on bone marrow examination and was found to be CD5 positive B-cell acute lymphoblastic leukemia on immunophenotyping. Cytogenetic analysis revealed translocation t(9:22).

Transfusion associated graft versus host disease.

This case report describes an immunocompetent lady who developed transfusion associated graft versus host disease following transfusion from a close relative. The diagnosis was established by HLA typing and STR analysis of the patient and her family.

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.

Microalbumiuria and associated risk factors in type II diabetics.

OBJECTIVE: To determine the frequency of microalbuminuria (MA) and its associated medical risk factors in type II diabetic patients. PLACE AND DURATION OF STUDY: This cross-sectional analytical study was conducted during Ist half of 2003 at Combined Military Hospital, Lahore. Non-probability purposive sampling was employed. MATERIALS AND METHODS: Study population included 150 type II diabetic patients (70 women, 80 men) attending outpatient department of the hospital. Patients having clinical albuminuria and with other causes of proteinuria were excluded. RESULTS: Women and men were of comparable ages. Women (26.4 kg/m2) had higher body mass index (BMI) than men (24.3 kg/m2). The frequency of MA was 46.7%, higher in males (50.6%) than females (41.5%). Fasting plasma glucose and HbA1c levels were significantly higher in patients with MA compared to those with normoalbuminuria (p <0.001). The microalbuminuric patients had significantly decreased HDL-c levels compared to normoalbuminuric subjects (p < 0.001). However, no relation of MA with age, gender, known duration of diabetes, BMI, history of smoking, hypertension and serum: total cholesterol, LDL - c, triglyceride, urea and creatinine was found. CONCLUSION: There is a strong association of poor glycaemic control and decreased HDL-c levels with the presence of microalbuminuria.