Rituximab-treated patients with lymphoma develop strong CD8 T-cell responses following COVID-19 vaccination.

B-cell depletion induced by anti-cluster of differentiation 20 (CD20) monoclonal antibody (mAb) therapy of patients with lymphoma is expected to impair humoral responses to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) vaccination, but effects on CD8 T-cell responses are unknown. Here, we investigated humoral and CD8 T-cell responses following two vaccinations in patients with lymphoma undergoing anti-CD20-mAb therapy as single agent or in combination with chemotherapy or other anti-neoplastic agents during the last 9 months prior to inclusion, and in healthy age-matched blood donors. Antibody measurements showed that seven of 110 patients had antibodies to the receptor-binding domain of the SARS-CoV-2 Spike protein 3-6 weeks after the second dose of vaccination. Peripheral blood CD8 T-cell responses against prevalent human leucocyte antigen (HLA) class I SARS-CoV-2 epitopes were determined by peptide-HLA multimer analysis. Strong CD8 T-cell responses were observed in samples from 20/29 patients (69%) and 12/16 (75%) controls, with similar median response magnitudes in the groups and some of the strongest responses observed in patients. We conclude that despite the absence of humoral immune responses in fully SARS-CoV-2-vaccinated, anti-CD20-treated patients with lymphoma, their CD8 T-cell responses reach similar frequencies and magnitudes as for controls. Patients with lymphoma on B-cell depleting therapies are thus likely to benefit from current coronavirus disease 2019 (COVID-19) vaccines, and development of vaccines aimed at eliciting T-cell responses to non-Spike epitopes might provide improved protection.

Influence of Malondialdehyde and Matrix Metalloproteinase-9 on Progression of Carotid Atherosclerosis in Chronic Renal Disease with Cardiometabolic Syndrome.

Objective was to assess whether the concentration of malondialdehyde (MDA) as a marker of lipid peroxidation and serum concentration of matrix metalloproteinase-9 (MMP-9) are involved in the process of atherosclerosis in chronic kidney disease (CKD) patients nondialysis-dependent and those on peritoneal dialysis (PD), both with signs of cardiometabolic syndrome (CMS). Thirty CKD and 22 PD patients were included in a study. All observed patients were divided into three subgroups depending on the degree of atherosclerotic changes in the carotid arteries (CA). Severity of atherosclerotic changes in the CA was evaluated by ultrasonography. We confirmed significantly lower level of serum MDA throughout all the stages of atherosclerosis in PD patients compared with observed CKD patients (P < 0.05) and increased serum concentration of MDA and MMP-9 with the progression of severity atherosclerotic changes in both groups of patients. The multiple regression analysis revealed that MDA and MMP-9 are significant predictors of changes in IMT-CA CKD patients (P < 0.05) and plaque score on CA in these patients (P < 0.05). The results suggest that MDA and MMP-9 could be mediators of CKD-related vascular remodeling in CMS.

Detection methods of immunoglobulin IgG in CSF and serum.

INTRODUCTION: This study represents a new approach to the extended analysis of correlation of findings of oligoclonal bands on gels and the level of intrathecal synthesis of immunoglobulin G in the central nervous system. Previous studies have shown that there is no correlation at this level as well as the number of tape or finding does not correlate with the forecast effect of therapy or patient outcome. AIMS OF THE STUDY: To determine the correlation of level of immunoglobulins IgG in CSF with the number of oligoclonal bands on the gel. MATERIAL AND METHODS: The retrospective study based on data processed in Clinical Immunology Clinical Center University of Sarajevo. Patients were assumed of multiple sclerosis according to clinical findings and magnetic resonance imaging. All CSF and serum samples were processed by nephelometry, isoelectric focusing on the gel. Statistical analysis of results was also performed by using SPSS statistical analysis program. RESULTS: Analyses were performed on 254 samples of cerebrospinal fluid and serum of patients from neurological clinic, suspected of multiple sclerosis. We concluded that there is no correlation between the level of intrathecal synthesis obtained by Reibergram with the number of oligoclonal bands on gels. We think that the reason could be a small sample of patients analyzed and it leaves room for future analysis on a larger sample. DISCUSSION AND CONCLUSION: For most patients with established MS we found intrathecal humoral response, type two, and the number and arrangement of IgG bands generally does not change during the disease, because they reflect long-term non-specific immune stimulation rather than a specific immune response that during infectious disease changes (quantitatively and qualitatively).

Prevalent and immunodominant CD8 T cell epitopes are conserved in SARS-CoV-2 variants.

The emergence of SARS-CoV-2 variants of concern (VOC) is driven by mutations that mediate escape from neutralizing antibodies. There is also evidence that mutations can cause loss of T cell epitopes. However, studies on viral escape from T cell immunity have been hampered by uncertain estimates of epitope prevalence. Here, we map and quantify CD8 T cell responses to SARS-CoV-2-specific minimal epitopes in blood drawn from April to June 2020 from 83 COVID-19 convalescents. Among 37 HLA ligands eluted from five prevalent alleles and an additional 86 predicted binders, we identify 29 epitopes with an immunoprevalence ranging from 3% to 100% among individuals expressing the relevant HLA allele. Mutations in VOC are reported in 10.3% of the epitopes, while 20.6% of the non-immunogenic peptides are mutated in VOC. The nine most prevalent epitopes are conserved in VOC. Thus, comprehensive mapping of epitope prevalence does not provide evidence that mutations in VOC are driven by escape of T cell immunity.

A T cell receptor targeting a recurrent driver mutation in FLT3 mediates elimination of primary human acute myeloid leukemia in vivo.

Acute myeloid leukemia (AML), the most frequent leukemia in adults, is driven by recurrent somatically acquired genetic lesions in a restricted number of genes. Treatment with tyrosine kinase inhibitors has demonstrated that targeting of prevalent FMS-related receptor tyrosine kinase 3 (FLT3) gain-of-function mutations can provide significant survival benefits for patients, although the efficacy of FLT3 inhibitors in eliminating FLT3-mutated clones is variable. We identified a T cell receptor (TCR) reactive to the recurrent D835Y driver mutation in the FLT3 tyrosine kinase domain (TCRFLT3D/Y). TCRFLT3D/Y-redirected T cells selectively eliminated primary human AML cells harboring the FLT3D835Y mutation in vitro and in vivo. TCRFLT3D/Y cells rejected both CD34+ and CD34- AML in mice engrafted with primary leukemia from patients, reaching minimal residual disease-negative levels, and eliminated primary CD34+ AML leukemia-propagating cells in vivo. Thus, T cells targeting a single shared mutation can provide efficient immunotherapy toward selective elimination of clonally involved primary AML cells in vivo.

Monitoring of disease biomarkers activity and immunophenotyping as important factors in SLE clinical management.

The highly specific biomarkers for monitoring of SLE disease activity are not yet defined up to date, due to existing of different clinical SLE phenotypes caused by individual genetic variation. Basically, numerous clinical complications follow SLE patients such as nephritis, atherosclerosis and cardial, CNS, gastrointestinal and ophthalmological complications, as well. Their monitoring in clinical SLE management can be evaluated by analysing of specific biochemical parameters and require permanent clinical observation. The presence of ANAs and anti-ds-DNAs are usual diagnostic SLE autoimmunity parameters, while SLE disease activity biomarkers are C3 and C4 level, anticardiolipin antibodies, anti-Sm/RNPs and, recently level of CD4 and CD8 lymphocytes. However, the number of TCR molecules on the T-cells surface at SLE patients is lower then in normal condition, and otherwise for these receptors CD molecules make specific connection. On the other hand, the T lymphocytes can be also, therapeutical targets at SLE patients, because of their clear direct involving in SLE pathogenesis. The SLE phenotypes are characterized by double CD negativity ( CD3 +/-, CD4-) caused by abnormal level of IL-2 and IL-17. T-lymphocytes have usually alpha-beta and gamma-delta TCR receptors, but for SLE patients is characteristic lower number gama-delta TCR molecules, detected in the peripheral blood specimens. Taking into account all of the facts, we investigated the level of specific usual SLE activity biomarkers (anti-ds-DNAs, C3, C4, anticardiolipin antibodies (beta-2-IgG, beta-2-IgM, ACA-G, ACA-M, CD4 and CD8 level) in serum specimens of SLE patients who underwent to the corresponding chemotherapy in combination with other biochemical and clinical parameters. Once again proved to be, that SLE biomarker monitoring, could be useful aproach for SLE activity disease and prediction organ damage, as well. In our investigation we used the following methods: immunofluorescence microscopy (IFA-ANA), and nephelometry, Hycor ELISA system and Flow cytometry, for precisely quantitative measurements. We determined correlation between C3 and C4 complement components level, CD3 (T-Ly), CD3+/HLA-DR and total HLA-DR with regard to SLE disease activity. Also, CD4 (Th), CD4:CD8 ratio, beta-2-G, beta-2-M not proved to be useful biomarkers in this sense, despite some results specific for some special SLE phenotypes. Anti-Sm/ RNPs proved to be better in SLE diagnostic process.

Reibergram and oligoclonal bands in diagnosis of multiple sclerosis.

INTRODUCTION: In this study authors have analyzed the correlation between the IgG immunoglobulins in cerebrospinal fluid and the findings of oligoclonal bands on gel. Immunoglobulin IgG in cerebrospinal fluid (CSF) can be detected in neurological diseasses (infections and inflammatory neurological diseases and in demyelinating diseases, like multiple sclerosis (MS)). Quantitative IgG in CSF can be expressed by different formulae Reiber (Reiber and Felgenhauer 1987), Tourtellotte (Tourtellotte 1970), Schuller (Schuller and Sagar 1983) and IgG Index (Link and Tibbling 1977). In this study we used Reibergram. Qualitative CSF IgG can be measured by electrophoresis and isoelectric focusing (IEF). We used IEF for analysig CSF and seum because of its higher sensitivity. AIMS OF THE STUDY: To determine the correlation of immunoglobulins IgG positivity in CSF with the finding of oligoclonal bands on the gel. MATERIAL AND METHODS: The retrospective study based on data processed in OJ Clinical Immunology KCUS. Patients were suspicious of multiple sclerosis according to clinical findings and magnetic resonance imaging. All CSF and serum samples were processed by nephelometry, isoelectric focusing on the gel. Statistical analysis of intrathecal synthesis was also performed according to Reibergram. RESULTS: Analyses were performed on 76 samples of cerebrospinal fluid and serum of patients from neurological clinic, suspected of multiple sclerosis. We received following results: 42 samples tested had type 1.25 samples tested showed type 2.3 samples had type 3.5 samples had type 4.1 sample had a fifth type. When we compare these results with values obtained by intrathecal synthesis of which is determined by Reibergram we obtained the following values: 16 samples had intrathecal synthesis of 20%-60%, 9 samples had a negative value of intrathecal synthesis of 10% or less. DISCUSSION AND CONCLUSION: For most patients with established MS we found intrathecal humoral response, type two, and the number and arrangement of IgG bands generally does not change during the disease, because they reflect long-term non-specific immune stimulation rather than a specific immune response that during infectious disease changes (quantitatively and qualitatively).

Correlation of autoantibodies presence detected by IFA-anti-dsDNA, IFA-AMA and immunoblotting with corresponding data in clinical management of autoimmune diseases.

Diagnosis and management of patients with SLE (Systemic Lupus Eritematosus), autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), involves specific diagnostic tests, such as IFA-AMA, IFA anti-dsDNA and immunoblotting for the detection of autoantibodies for specific autoantigens (mitochondria, dsDNA, M2, LKM-1, LC-1, SLA/LP). We established specific correlation between the detected autoantibodies and corresponding clinical findings. The total of 813 serum specimens were probed with IFA-anti-dsDNA, 98 of which tested positive. We also performed dilution analysis to the end point for all the positive specimens. Numerous specimens were tested by IFA, AMA and immunoblotting.

Prognostic indicators of adverse renal outcome and death in acute kidney injury hospital survivors.

INTRODUCTION: Data regarding prognostic factors of post-discharge mortality and adverse renal function outcome in acute kidney injury (AKI) hospital survivors are scarce and controversial. OBJECTIVES: We aimed to identify predictors of post-discharge mortality and adverse renal function outcome in AKI hospital survivors. PATIENTS AND METHODS: The study group consisted of 84 AKI hospital survivors admitted to the tertiary medical center during 2-year period. Baseline clinical parameters, with renal outcome 3 months after discharge and 6-month mortality were evaluated. According survival and renal function outcome, patients were divided into two groups. RESULTS: Patients who did not recover renal function were statistically significantly older (P < 0.007) with higher Charlson comorbidity index (CCI) score (P < 0.000) and more likely to have anuria and oliguria (P = 0.008) compared to those with recovery. Deceased AKI patients were statistically significantly older (P < 0.000), with higher CCI score (P < 0.000), greater prevalence of sepsis (P =0.004), higher levels of C-reactive protein (CRP) (P < 0.017) and ferritin (P < 0.051) and lower concentrations of albumin (P<0.01) compared to survivors. By multivariate analysis, independent predictors of adverse renal outcome were female gender (P =0.033), increasing CCI (P =0.000), presence of pre-existing chronic kidney disease (P =0.000) and diabetes mellitus (P =0.019) as well as acute decompensated heart failure (ADHF) (P =0.032), while protective factor for renal function outcome was higher urine output (P =0.009). Independent predictors of post-discharge mortality were female gender (P =0.04), higher CCI score (P =0.001) and sepsis (P =0.034). CONCLUSION: Female AKI hospital survivors with increasing burden of comorbidities, diagnosis of sepsis and ADHF seem to be at high-risk for poor post-discharge outcome.

ELISA Test for Analyzing of Incidence of Type 1 Diabetes Autoantibodies (GAD and IA2) in Children and Adolescents.

INTRODUCTION: Anti GAD (antibodies on glutamic acid decarboxylase) and anti-IA2 antibodies (against tyrosine phosphatase), today, have their place and importance in diagnosis and prognosis of Type 1 diabetes. Huge number of patients with diabetes mellitus type 1 have these antibodies. Insulin antibodies are of critical importance in diagnosis of diabetes mellitus type 1 for pediatric population. MATERIALS AND METHODS: During 2014, the samples of 80 patients from Clinical Center University Sarajevo (CCUS) Pediatrics clinic's, Endocrinology department were analyzed on anti-GAD and IA2 antibodies. The samples of serums of all patients were analyzed with ELISA tests using Anti GAD ELISA (IgG) kites from EUROIMMUN company. These are quantitative in vitro tests for human antibodies against decarboxylase of glutamine acid (GAD) and IA2, in serum or EDTA plasm. RESULTS: During the period of one year, in CCUS's Organizational unit, Institute for Clinical Immunology, 80 samples of patients with anti GAD and IA2 antibodies were analyzed. Out of total number of samples, 41 were male patients, or 51% and 39 female, or 49%. The youngest patient was born in 2012, and the oldest in 1993. Age average was represented by the patients born in 2001. Share of positive results for IA2 antibodies and GAD antibodies was 37% for IA2 antibodies, and 63% for GAD antibodies. DISCUSSION: During an autoimmune - mediated Diabetes mellitus type 1 leads to T-cell mediated destruction of beta cells of pancreatic islets, reduced production of insulin and glucose metabolism. Studies have shown that these bodies are the most intense single marker for identifying persons with increased risk for diabetes development.

Assessments of the socioeconomic status and diet on the prevalence of dental caries at school children in central bosnian canton.

AIM: The main aim of this research was to determine the influence of socioeconomic status and residence/living conditions on the status of oral health (e.g. health of mouth and teeth) in primary school students residing in Canton Central Bosnia. METHODS: The study was designed as a cross-sectional study. Our research included two-phased stratified random sample of 804 participants. The quantitative research method and newly designed survey instrument were utilized in order to provide data on the oral health of the examined children. The alternate hypothesis foresaw that "there were significant statistical differences between the levels of incidence of dental caries in comparison to the incidence in children of different socioeconomic status. RESULTS: The Chi square () of 22.814, degree of freedom (Df) = 8, coefficient of contingency of 0.163 and T-test (Stat) of-0.18334 showed that there were no significant statistical differences at p < 0.05 level between the primary school children from urban and rural areas. The obtained results showed that the caries indexes in elementary schools in Central Bosnia Canton were fairly uniform. Research showed that there were a difference in the attitudes towards a regular dental visits, which correlated with social-educational structure of the children's' families. CONCLUSION: According to the results, we can see that the socioeconomic status of patients had an effect on the occurrence of dental caries and oral hygiene in patients in relation to the rural and urban areas, because we can see that by the number of respondents, the greater unemployment of parents in both, rural and urban areas, caused a host of other factors, which were, either, directly or indirectly connected with the development of caries.

Tumor necrosis factor-alpha serum level in assessment of disease activity in inflammatory bowel diseases.

AIM: To investigate an influence of the concentration of proinflammatory cytokines tumor necrosis factor-alpha (TNF-α) in serum on the activity of inflammatory bowel disease (IBD). METHODS: The IBD patients of both genders (n=60) were divided in two equal groups, ulcerative colitis (UC) and Crohn's disease (CD). Based on the result of activity index each group was subdivided in two subgroups: active and inactive phase of the disease. Age and gender matched apparently healthy individuals (n=30) involved in the control group. Serum TNF-α concentration was determined by enzyme linked immune-adsorbent assay (ELISA). RESULTS: The significant difference (Mann-Whitney Test) in serum TNF-α level was found between healthy controls 28.86 pg/ml (28.74 - 29.19 pg/ml) and CD patients (29.47 pg/ml (29.1 - 29.77 pg/ml) (p less than 0.05) and UC patients 29.34 pg/ml (29.14 - 29.71 pg/ ml) (p less than 0.05) respectively. Serum TNF-α level in patients with CD was higher compared to serum TNF-α level in patients with UC, but the difference was not significant (p more than 0,05). There were no significant difference in serum TNF-α concentrations either in CD or UC patients related to the phase of disease activity: active CD 29.53 pg/ml (29.20 - 29.90 pg/ml) vs inactive CD 29.26 pg/ml (29.15 - 29.53 pg/ml); active UC 29.53 pg/ml (29.32 - 29.85 pg/ ml) vs inactive UC 29.26 pg/ml (29.10 - 29.63 pg/ml). CONCLUSIONS: Since there were no differences in serum TNF-α concentrations related to the disease activity we consider that TNF-α is not an adequate serum biomarker for an assessment of the disease activity in patients with IBD.

ELISA subtypization of anti-ENA autoantibodies in clinical management of autoimmune diseases in Bosnia and Herzegovina.

The basis of autoimmune diseases such as SLE (Systemic Lupus Eritematodes), Sjogren's syndrome, scleroderma, dermatomyositis and polymiositis is the creation of auto-antibodies to the following specific extractable nuclear antigens (ENA):Jo-1, Ssl-70, SS-A, SS-B, Sm and Sm/RNPs. Some of these antigens are in fact enzymes (Jo-1-histidil-tRNA synthetase, Scl-70-topoisomerase) which are inhibited by specific autoantibodies--this leads to disturbance in the metabolism of DNA and protein biosynthesis. During 2009, we analyzed total of 87 serum samples of patients suspected for autoimmune disorder using ANA-IFA and ELISA-ENA-6 methods. After establishing IFA-ANA positivity (83.9%), all serum specimens; ANA positive and negative, were subtypized by ELISA ENA-6 test. Analysis showed the highest incidence of anti-SS-A (56%), and incidence of anti-SS-B (29.8%), anti-Sm/ RNP (11.5%), anti-Jo-1 (2.3%) and anti-Scl-70 (1,1%) auto-antibodies. Also, 78.5% of IFA-ANA negative serum specimens showed high level of positivity (212.50 and 277.0 IU/ml) to SS-A (78.5%) and SS-B (21.4%) antigenes using ELISA-ENA-6 subtypization. Following these results, we conclude that it is necessary to introduce Western blot confirmation testing. After comparing with other clinical findings, we diagnosed the following autoimmune diseases: SLE, Sjogren's syndrome and dermatomiosytis.

Transplantation of organs: one of the greatest achievements in history of medicine.

The history of transplantation is a scientific journey describing the medical community's effort to understand how the human body works. Humans have long realized the possibilities which transplantation of organs and tissue provides. Throughout history people have always been intrigued by the possibilities of the transplantation of organs and tissues. In the 6th Century BC Indian surgeons described how to reconstruct facial wounds by transplanting skin from one place on the body to the other. During the middle age there were many references in historical medical literature of attempted blood transfusions as well as the transplantation of teeth. A skin transplant and a corneal transplant were reported in medical journals dating as far back as 1880. These early attempts were usually unsuccessful. Early in the twentieth century transplantation started to offer the promise of restored health and life. One of the exceptional medical advances of the twentieth century, organ transplantation has become a routine treatment for patients with organ failure which was a goal.

Comparative study of interleukin 1ALFA and interleukin 6 concentrations in serum specimens detected by ELISA.

Interleukin 1 (IL-1) contains two proteins, which are the products of distinct genes, but which recognize the same cell surface receptors. In the liver, IL-1 initiates the acute phase response resulting in an increase in hepatic protein synthesis and decreased albumin production IL-1 also plays an important role in immune functions, having effects on macrophages/monocytes, T lymphocytes, B lymphocytes, NK cells, and LAK cells. Interleukin-6 (IL-6) is a cytokine that regulates immune responses. We analyzed total 160 serum specimens of patients from Clinical Center University of Sarajevo with different inflammatory diseases by ELISA method on interleukins: IL-1alfa and IL-6. Tests that we performed with IL-lalfa and IL-6 by ELISA method confirmed that serum specimens with IL-6 ELISA showed increased values of tested specimens, than the lowest standard and blank. We had average levels of IL-1alfa 3.7 pg/ml which was below the level of the lowest standard. All obtained results were in accordance with the results in IBL protocol for blank and lowest standard values, as well as the average levels of serum specimen values.