Clinical association with non-hodgkin lymphoma

Introduction: The association of anti phospholipid antibodies (APA) with Non..Hodgkin Lymphoma~) has been reported and complete remission was associated with disappearance of APA. APA are autoantibodies which include anticardiolipin antibody (ACA), anti-beta2-glycoprotein-I (~2 GPI) antibodies and lupus anticoagulant (LA). Objective: The purpose of this study was to determine the prevalence of APA and its clinical association among NHL patients in our institution. Meth~dology: A study was conducted ~n Hospital· Unive~sity Science Malaysia . . .. . (HUSM) between June 2003 and July 2004. Fifty-three selected NHL patients · were tested for ACA and anti-~2 GPI at presentation using ELISA technique. They were followed-u'p over a median period of 6 months to detect the occurrence of thromboembolism (TE} and bone marrow recovery following chemotherapy using full blood counts. Results: APA was found in 23 out of 53 NHL patients, with ACA 35.8o/o and anti( 32 GPI antibodies 18.9o/o. The incidence of elevated APA was increased after 40. years old (91.3°/o). However, positivity for APA was not associated with gender, survival, histology or stage of lymphoma. There were three patients who developed TE and all of them were ACA positive. Positive APA was found to correlate with thrombocytopenia at presentation (p= 0.032) and associated with poor platelet recovery following chemotherapy (p=0.001 ). Conclusion: APA was common among NHL patients especially after 40 years old. Although no statis~ical association was found between APA positive NHL and thrombosis, there was a tendency forTE to occur in APA positive NHL

A New Nested Allele-Specific Multiplex Polymerase Chain Reaction Method for Haplotyping of VKORC1 Gene to Predict Warfarin Sensitivity

The vitamin K epoxide reductase complex 1 gene (VKORC1) is commonly assessed to predict warfarin sensitivity. In this study, a new nested allele-specific multiplex polymerase chain reaction (PCR) method that can simultaneously identify single nucleotide polymorphisms (SNPs) at VKORC1 381, 861, 5808, and 9041 for haplotype analysiswas developed and validated. ExtractedDNAwas amplified in the first PCR DNA, which was optimized by investigating the effects of varying the primer concentrations, annealing temperature, magnesium chloride concentration, enzyme concentration, and the amount of DNA template. The amplification products produced from the first round of PCR were used as templates for a second PCR amplification in which both mutant and wild-type primers were added in separate PCR tubes, followed by optimization in a similar manner. The final PCR products were resolved by agarose gel electrophoresis and further analysed by using a VKORC1 genealogic tree to infer patient haplotypes. Fifty patients were identified to have H1H1, one had H1H2, one had H1H7, 31 had either H1H7 or H1H9, one had H1H9, eight had H7H7, and one had H8H9 haplotypes. This is the first method that is able to infer VKORC1 haplotypes using only conventional PCR methods

In Vitro

This study aimed to evaluate in vitro whole blood (WB) clot lysis method for the assessment of fibrinolytic activity. Standardized unresected (uncut) retracted WB clot was incubated in pool platelet poor plasma (PPP) for varying incubation times and in streptokinase (SK) at different concentrations. The fibrinolytic activity was assessed by D-dimer (DD), confocal microscopy, and clot weight. DD was measured photometrically by immunoturbidimetric method. There was a significant difference in mean DD levels according to SK concentrations (P=0.007). The mean DD±SD according to the SK concentrations of 5, 30, 50, and 100 IU/mL was: 0.69±0.12, 0.78±0.14, 1.04±0.14 and 2.40±1.09 μg/mL. There were no significant changes of clot weight at different SK concentrations. Gradual loss and increased branching of fibrin in both PPP and SK were observed. Quantitation of DD and morphology of fibrin loss as observed by the imaging features are in keeping with fibrinolytic activity. Combination of DD levels and confocal microscopic features was successfully applied to evaluate the in vitro WB clot lysis method described here

Performance Comparison Between Conventional Fluorescent Spot Test and Quantitative Assay in Detecting G6PD Deficiency in Neonates

Objectives: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy worldwide. The fluorescent spot test (FST) is the conventional method for screening neonates for G6PD. However, it has limitations and quantitative assays such as the CareStart Biosensor 1 are being increasingly recommended. This study aimed to compare FST and CareStart Bioensor 1 in their ability to detect G6PD levels in neonates. Methods: This cross-sectional study involved 455 neonates between June and December 2020. Two milliliters of cord blood were analyzed with CareStart Biosensor 1 and dried cord blood spots with FST. Data was recorded and statistically analyzed. Sensitivity, specificity, positive predictive value, and negative predictive value were calculated to determine the performance of FST at specific G6PD cut-off values; Cohen’s kappa analysis assessed the agreement between the two methods. Results: The sensitivity of FST at 30% cut-off G6PD activity level was 91.0%, (95% CI: 57.0–100) and specificity of 97.0% (95% CI: 95.0–98.0). At 60% cut-off, the FST sensitivity sharply declined to 29.0% (95% CI: 19.0–40.0) with a specificity of 100% (95% CI: 98.0–100). The overall prevalence of G6PD deficiency was 5.1% as measured by FST and 17.8% by Biosensor 1 (p< 0.001). Conclusions: In this study, FST missed a significant proportion of cases of intermediate G6PD levels. FST also misclassified several G6PD intermediate individuals as normal, rendering them susceptible to oxidative stress. Biosensor 1 reported a significantly higher prevalence of G6PD deficiency

Epigenetic insights and potential modifiers as therapeutic targets in β–thalassemia

Thalassemia, an inherited quantitative globin disorder, consists of two types, α– and β–thalassemia. β–thalassemia is a heterogeneous disease that can be asymptomatic, mild, or even severe. Considerable research has focused on investigating its underlying etiology. These studies found that DNA hypomethylation in the β–globin gene cluster is significantly related to fetal hemoglobin (HbF) elevation. Histone modification reactivates γ-globin gene expression in adults and increases β–globin expression. Down-regulation of γ–globin suppressor genes, i.e., BCL11A, KLF1, HBG-XMN1, HBS1L-MYB, and SOX6, elevates the HbF level. β–thalassemia severity is predictable through FLT1, ARG2, NOS2A, and MAP3K5 gene expression. NOS2A and MAP3K5 may predict the β–thalassemia patient’s response to hydroxyurea, a HbF-inducing drug. The transcription factors NRF2 and BACH1 work with antioxidant enzymes, i.e., PRDX1, PRDX2, TRX1, and SOD1, to protect erythrocytes from oxidative damage, thus increasing their lifespan. A single β–thalassemia-causing mutation can result in different phenotypes, and these are predictable by IGSF4 and LARP2 methylation as well as long non-coding RNA expression levels. Finally, the coinheritance of β–thalassemia with α–thalassemia ameliorates the β–thalassemia clinical presentation. In conclusion, the management of β–thalassemia is currently limited to genetic and epigenetic approaches, and numerous factors should be further explored in the future

The first Malay database toward the ethnic-specific target molecular variation

BACKGROUND:The Malaysian Node of the Human Variome Project (MyHVP) is one of the eighteen official Human Variome Project (HVP) country-specific nodes. Since its inception in 9(th) October 2010, MyHVP has attracted the significant number of Malaysian clinicians and researchers to participate and contribute their data to this project. MyHVP also act as the center of coordination for genotypic and phenotypic variation studies of the Malaysian population. A specialized database was developed to store and manage the data based on genetic variations which also associated with health and disease of Malaysian ethnic groups. This ethnic-specific database is called the Malaysian Node of the Human Variome Project database (MyHVPDb). FINDINGS:Currently, MyHVPDb provides only information about the genetic variations and mutations found in the Malays. In the near future, it will expand for the other Malaysian ethnics as well. The data sets are specified based on diseases or genetic mutation types which have three main subcategories: Single Nucleotide Polymorphism (SNP), Copy Number Variation (CNV) followed by the mutations which code for the common diseases among Malaysians. MyHVPDb has been open to the local researchers, academicians and students through the registration at the portal of MyHVP ( http://hvpmalaysia.kk.usm.my/mhgvc/index.php?id=register ). CONCLUSIONS:This database would be useful for clinicians and researchers who are interested in doing a study on genomics population and genetic diseases in order to obtain up-to-date and accurate information regarding the population-specific variations and also useful for those in countries with similar ethnic background

A comparative study on fibrinolytic in acute stroke patients in HUSM

The fibrinolytic system plays an important role in preventing intra-vascular thrombosis. Previous study bad shown the pathogenesis of stroke is related to abnormality in fibrinolytic system. This study was conducted in HUSM for a one-year period from March 2005 to February 2006. Stroke patients were selected from adult wards whereas control individuals were chosen from various clinics. This study was done to compare the levels of three fibrinolytic markers i.e. plasminogen (plg), tissue-plasminogen activator (t-PA) and plasminogen activator inhibitor type-I (PAI-l) between acute stroke patients and stable non-stroke individuals and to investigate the clinical significance of these markers. One hundred and six individuals and 51 acute stroke patients were selected. Both groups have similar risk factors. Their bloods were tested for the level of t-PA and PAl-l using EUSA technique (Biopool TintElize) whereas plasminogen was tested with colorimetric -··· . -· assay using Hemosil Tm. They were follow-up over a period of 3 months to detect their survival and lec;overy. We found only t-PA level was significantly higher in acute stro.k e patients compared to . control group even after adjusting the cofo~ders using ANCOV A test. Plasminogen and PAI-l showed no significant statistical association between both groups. For PAI-l, the mean level is lower in stroke group after adjusting the confounders. There are no significant statistical association between the three fibrinolytic markers and age, gender, manber of risk factors, disease severity, survival and neurological recovery. We observed all the eight patients who died dwing hospitalization or at the time of follow-up possessed high level oft-PA although statistically not significant. The limitation of this study was that we could not show the abnormality in t-PA is primary or secondary events to the development of stroke. High t-PA level indicates abnormal intravascular fibrinolysis which is probably an initiator ~f the-eerebrevaseular--- .. event or indicating of underlying vascular damage. This finding supports the hypothesis that disturbances in fibrinolysis occur in stroke patients during cerebrovascular event. We found an association between high t-PA antigen level and stroke with a 4.6-fold odd ratio. t-PA level could be a marker to predict high-risk patients for stroke development. Using haemOstatic model addressing fibrinolytic system is a potential discovery for future therapy to prevent acute episodes of stroke together with other therapeutic agents

von Willebrand Factor Propeptide: A Potential Disease Biomarker Not Affected by ABO Blood Groups

Epidemiological studies have shown that vascular-related disorders are associated with high von Willebrand factor antigen (VWF:Ag) and VWF propeptide (VWFpp). VWFpp is secreted together with VWF:Ag upon endothelial cell activation, hence it could be a potential biomarker. This study was conducted to compare between VWFAg and VWFpp levels among 30 healthy individuals and 42 patients with high levels of VWF:Ag in different medical conditions and ABO blood groups. VWFpp levels were strongly correlated with VWF:Ag. VWF:Ag and VWFpp levels were significantly increased in patients compared to healthy individuals. VWFpp is not affected by ABO blood group in both healthy individual and patient groups unlike VWF:Ag. As expected, this study showed that VWFpp levels increased in parallel with VWF:Ag levels in patients with diseases associated with endothelial activation. VWFpp though nonspecific is a potential biomarker reflecting underlying pathophysiological changes in various medical conditions with an additional advantage of not being influenced by ABO blood groups

Conventional Rapid Latex Agglutination in Estimation of von Willebrand Factor: Method Revisited and Potential Clinical Applications

Measurement of von Willebrand factor antigen (VWF : Ag) levels is usually performed in a specialised laboratory which limits its application in routine clinical practice. So far, no commercial rapid test kit is available for VWF : Ag estimation. This paper discusses the technical aspect of latex agglutination method which was established to suit the purpose of estimating von Willebrand factor (VWF) levels in the plasma sample. The latex agglutination test can be performed qualitatively and semiquantitatively. Reproducibility, stability, linearity, limit of detection, interference, and method comparison studies were conducted to evaluate the performance of this test. Semiquantitative latex agglutination test was strongly correlated with the reference immunoturbidimetric assay (Spearman’s rho = 0.946, P<0.001, n=132). A substantial agreement (κ=0.77) was found between qualitative latex agglutination test and the reference assay. Using the scoring system for the rapid latex test, no agglutination is with 0% VWF : Ag (control negative), 1+ reaction is equivalent to <20% VWF : Ag, and 4+ reaction indicates >150% VWF : Ag (when comparing with immunoturbidimetric assay). The findings from evaluation studies suggest that latex agglutination method is suitable to be used as a rapid test kit for the estimation of VWF : Ag levels in various clinical conditions associated with high levels and low levels of VWF : Ag