Publications by authors named "Meganathan Kannan"

36 Publications

Study of Prothrombotic Gene Variations Associated with the Risk of Development of Thrombosis in Patients with Down Syndrome.

Indian J Hematol Blood Transfus 2021 Jul 17;37(3):507-508. Epub 2020 Oct 17.

Division of Blood and Vascular Biology, Department of Life Sciences, School of Life Sciences, Central University of Tamil Nadu, Thiruvarur, 610101 India.

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http://dx.doi.org/10.1007/s12288-020-01369-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8239101PMC
July 2021

The COVID-19 Pandemic and the Need for an Integrated and Equitable Approach: An International Expert Consensus Paper.

Thromb Haemost 2021 Aug 24;121(8):992-1007. Epub 2021 Jun 24.

Division of Blood and Vascular Biology, Department of Life Sciences, School of Life Sciences, Central University of Tamil Nadu, Thiruvarur, India.

Background:  One year after the declaration of the coronavirus disease 2019 (COVID-19) pandemic by the World Health Organization (WHO) and despite the implementation of mandatory physical barriers and social distancing, humanity remains challenged by a long-lasting and devastating public health crisis.

Management:  Non-pharmacological interventions (NPIs) are efficient mitigation strategies. The success of these NPIs is dependent on the approval and commitment of the population. The launch of a mass vaccination program in many countries in late December 2020 with mRNA vaccines, adenovirus-based vaccines, and inactivated virus vaccines has generated hope for the end of the pandemic.

Current Issues:  The continuous appearance of new pathogenic viral strains and the ability of vaccines to prevent infection and transmission raise important concerns as we try to achieve community immunity against severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) and its variants. The need of a second and even third generation of vaccines has already been acknowledged by the WHO and governments.

Perspectives:  There is a critical and urgent need for a balanced and integrated strategy for the management of the COVID-19 outbreaks organized on three axes: (1) revention of the SARS-CoV-2 infection, (2) and early diagnosis of patients at risk of disease worsening, and (3) of medical care (PDA).

Conclusion:  The "PDA strategy" integrated into state policy for the support and expansion of health systems and introduction of digital organizations (i.e., telemedicine, e-Health, artificial intelligence, and machine-learning technology) is of major importance for the preservation of citizens' health and life world-wide.
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http://dx.doi.org/10.1055/a-1535-8807DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8322591PMC
August 2021

Association of VEGF and p53 Polymorphisms and Spiral Artery Remodeling in Recurrent Pregnancy Loss: A Systematic Review and Meta-Analysis.

Thromb Haemost 2021 May 26. Epub 2021 May 26.

Division of Blood and Vascular Biology, Department of Life Sciences, School of Life Sciences, Central University of Tamil Nadu, Thiruvarur, India.

Many studies have reported the association of -1154G/A, 936C/T, and Arg72Pro polymorphisms with recurrent pregnancy loss (RPL), but the outcomes are inconsistent. We have used a meta-analysis to associate these polymorphisms with RPL, having the spiral artery remodeling as a major risk factor. The studies were identified from three different reputed databases, namely ScienceDirect, PubMed/Medline, and Scopus. The eligible studies of 1154G/A, 936C/T, and Arg72Pro polymorphisms associated with the RPL were selected for the analysis. They were segregated into three different ethnic groups as Asians, Caucasians, and mixed population. For the analysis, the overall prevalence, odds ratio, risk ratio, relative risk ratio, and -values were calculated. A total of 3,241 RPL cases and 3,205 healthy controls from 21 different case-control studies were analyzed. RPL was highly prevalent in the mixed population with 1154G/A and Arg72Pro polymorphisms (70.04 and 66.46%, respectively) and in the Asian population with VEGF 936C/T polymorphism (53.58%). The homozygous recessive genotypes of and p53 exhibited significant association between the respective polymorphisms and RPL along with the increased risk of outcome. The current analysis conclusively reports the geographic distribution of the different genetic polymorphisms which shows high association with the progression of RPL. Understanding the spectrum of polymorphisms on different populations with the spiral artery remodeling as a risk factor encloses the importance of the vasculature during the pregnancy.
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http://dx.doi.org/10.1055/a-1518-1756DOI Listing
May 2021

The ISTH bleeding assessment tool as predictor of bleeding events in inherited platelet disorders: Communication from the ISTH SSC Subcommittee on Platelet Physiology.

J Thromb Haemost 2021 05;19(5):1364-1371

Department of Internal Medicine, IRCCS Policlinico S. Matteo Foundation, University of Pavia, Pavia, Italy.

Background: The ISTH Bleeding Assessment Tool (ISTH-BAT) has been validated for clinical screening of suspected von Willebrand disease (VWD) and for bleeding prediction. Recently it has been validated for subjects with inherited platelet disorders (IPD) (BAT-VAL study).

Objectives: To determine whether the ISTH-BAT bleeding score (BS) predicts subsequent bleeding events requiring treatment in IPD patients.

Methods: Patients with IPD, type 1 VWD (VWD-1) and age- and sex-matched healthy controls enrolled in the BAT-VAL study were prospectively followed-up for 2 years and bleeding episodes requiring treatment were recorded.

Results: Of the 1098 subjects initially enrolled, 955 were followed-up and 124 suffered hemorrhages during follow-up, 60% of whom had inherited platelet function disorders (IPFD). Total number of events was significantly higher in IPFD (n = 235) than VWD-1 (n = 52) or inherited thrombocytopenia (IT; n = 20). Events requiring transfusions were 66% in IPFD, 5.7% in VWD-1, and 3% in IT. Baseline BS was significantly higher in IPFD patients with a bleeding event at follow-up than in those without (p < .01) and the percentage of subjects suffering a bleeding event increased proportionally to baseline BS quartile. A significant association between the BS and the chance of suffering severe bleeding was found in the overall, IPFD, and VWD-1 populations. Similar results were obtained for the pediatric population.

Conclusions: Inherited platelet function disorder patients with high BS at enrollment are more likely to suffer from bleeding events requiring treatment at follow-up. Moreover, the higher the baseline BS quartile the greater the incidence of subsequent events, suggesting that independently from diagnosis a high BS is associated with a greater risk of subsequent hemorrhage.
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http://dx.doi.org/10.1111/jth.15263DOI Listing
May 2021

Increase of Plasma TNF- Is Associated with Decreased Levels of Blood Platelets in Clinical Dengue Infection.

Viral Immunol 2020 Jan/Feb;33(1):54-60. Epub 2019 Sep 18.

Division of Infection Biology, Department of Life Sciences, School of Life Sciences, Central University of Tamil Nadu, Thiruvarur, India.

Dengue virus (DENV) infection has become an increasingly common concern in tropical and subtropical regions. It has protean manifestations ranging from febrile phase to severe life-threatening illness. In this study, we estimated Th1 and Th2 cytokines and correlated the levels with dengue severity along with certain hematological and biochemical parameters. We also studied the seroprevalence of dengue between October and December 2017 at the Government Theni Medical College, India. Individuals with dengue fever (DF) were positive for either IgM or IgG, or both. The biochemical and hematological parameters along with plasma tumor necrosis factor alpha (TNF-), interferon-gamma (IFN-), granulocyte monocyte-colony stimulating factor (GM-CSF), interleukin (IL)-13, IL-12p70, IL-10, IL-5, IL-4, and IL-2 cytokines were estimated. The prevalence of DF was 42.9% during the study period. IL-2, TNF-, IL-4, and IL-10 levels were significantly elevated ( < 0.005) in patients with secondary DENV infection, whereas the level of IL-13 remained unaltered during both primary and secondary infections. No statistically significant difference was noticed with IL-12p70, IL-5, IFN-, and GM-CSF between the healthy controls and the primary and secondary DENV-infected groups. Increase of 1 unit of TNF- was associated with a decrease of 160 units of blood platelets. Together, the study suggests that TNF- could play a key role in the pathogenesis of dengue, and despite the decrease in platelet levels, it remains to be seen whether any other inflammatory cells regulate the levels of TNF- in DENV infection.
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http://dx.doi.org/10.1089/vim.2019.0100DOI Listing
February 2021

Platelet activation markers in evaluation of thrombotic risk factors in various clinical settings.

Blood Rev 2019 09 22;37:100583. Epub 2019 May 22.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Platelets play a major role in primary hemostasis and thrombus formation. After vascular injury, platelets adhere to injured site and rapidly change their shape that switches the resting platelets to active state. Activated platelets aggregate and secrete biologically active intermediate substances that further potentiate platelet activation through autocrine as well as paracrine mechanisms. The activated platelet expresses certain proteins that are not seen on the resting platelets, thus these proteins serve as markers of platelet activation. Other subsequent events of platelet activation include release of microvesicles and formation of complexes with other circulating cells, like monocytes and neutrophils. Platelet activation markers are useful tools in evaluating risk factors of thrombosis in a variety of clinical conditions. Increased platelet activation has been associated with various pathological conditions such as acute coronary syndrome, stroke, peripheral vascular disease and other inflammatory diseases. The advancement in technologies helps in determining the status of platelet activation in such clinical conditions. This article focuses on the sources, mechanism and diagnosis of platelet activation and their clinical implications.
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http://dx.doi.org/10.1016/j.blre.2019.05.007DOI Listing
September 2019

Characterization of VWF gene conversions causing von Willebrand disease.

Br J Haematol 2019 03 29;184(5):817-825. Epub 2018 Nov 29.

Department of Pediatric Haematology and Oncology, University Medical Centre, Eppendorf, Hamburg, Germany.

We previously reported that von Willebrand Factor gene (VWF) conversions are a relatively frequent cause of von Willebrand disease (VWD), however, their molecular pathomechanisms resulting in variant phenotypes is largely unknown. Here, we characterized VWF conversions harbouring missense and synonymous mutations, through generating a series of mutant constructs followed by transient expression in 293 cells, and qualitative and quantitative analysis of recombinant VWF (rVWF). The characterization of mutant rVWF showed the critical roles of synonymous variants in the pathogenicity of VWF conversions. The gene conversion variants p.Val1229Gly, p.Asn1231Thr, p.Asn1231Ser and p.Ala1464Pro in the absence of synonymous p.Ser1263= and p.Gln1449= showed minimal effect on rVWF synthesis and activity. Interestingly, a construct including the synonymous variants displayed significantly low rVWF expression and activity. The variant p.Pro1266Leu showed gain of rVWF function toward glycoprotein Ibα; surprisingly, this function was significantly abolished in the presence of gene conversion variants p.Val1229Gly-p.Asn1231Thr. Taken together, our expression studies suggest that synonymous variants in the combination of other gene conversion variants suppress the protein expression, possibly due to defective primary mRNA structure or processing. The variants p.Val1229Gly-p.Asn1231Thr affected the VWF gain of function caused by variant p.Pro1266Leu, probably due to conformational changes in VWF.
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http://dx.doi.org/10.1111/bjh.15709DOI Listing
March 2019

Viral Persistence and Chronicity in Hepatitis C Virus Infection: Role of T-Cell Apoptosis, Senescence and Exhaustion.

Cells 2018 Oct 12;7(10). Epub 2018 Oct 12.

Division of Infection Biology and Medical Microbiology, Department of Life Sciences, Central University of Tamil Nadu (CUTN), Thiruvarur 610005, India.

Hepatitis C virus (HCV) represents a challenging global health threat to ~200 million infected individuals. Clinical data suggest that only ~10⁻15% of acutely HCV-infected individuals will achieve spontaneous viral clearance despite exuberant virus-specific immune responses, which is largely attributed to difficulties in recognizing the pathognomonic symptoms during the initial stages of exposure to the virus. Given the paucity of a suitable small animal model, it is also equally challenging to study the early phases of viral establishment. Further, the host factors contributing to HCV chronicity in a vast majority of acutely HCV-infected individuals largely remain unexplored. The last few years have witnessed a surge in studies showing that HCV adopts myriad mechanisms to disconcert virus-specific immune responses in the host to establish persistence, which includes, but is not limited to viral escape mutations, viral growth at privileged sites, and antagonism. Here we discuss a few hitherto poorly explained mechanisms employed by HCV that are believed to lead to chronicity in infected individuals. A better understanding of these mechanisms would aid the design of improved therapeutic targets against viral establishment in susceptible individuals.
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http://dx.doi.org/10.3390/cells7100165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6210370PMC
October 2018

Phenotypic and molecular characterisation of type 3 von Willebrand disease in a cohort of Indian patients.

Thromb Haemost 2013 Apr 14;109(4):652-60. Epub 2013 Feb 14.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, 110029, India.

Severe type 3 VWD (VWD3) is characterised by complete absence or presence of trace amounts of non-functional von Willebrand factor (VWF). The study was designed to evaluate the VWF mutations in VWD3 patients and characterise the breakpoints of two identified homozygous novel large deletions. Patients were diagnosed by conventional tests and VWF multimer analysis. Mutation screening was performed in 19 VWD3 patients by direct sequencing of VWF including flanking intronic sequence and multiplex ligation-dependent probe amplification (MLPA) analysis. Breakpoint characterisation of two identified novel large deletions was done using walking primers and long spanning PCR. A total of 21 different mutations including 15 (71.4%) novel ones were identified in 17 (89.5%) patients. Of these mutations, five (23.8%) were nonsense (p.R1659*, p.R1779*, p.R1853*, p.Q2470*, p.Q2520*), one was a putative splice site (p.M814I) and seven (33.3%) were deletions (p.L254fs*48, p.C849fs*60, p.L1871fs*6, p.E2720fs*24) including three novel large deletions of exon 14-15, 80,830bp (-41510_657+7928A*del) and 2,231bp [1534-2072T_c.1692G*del(p.512fs*terminus)] respectively. A patient carried gene conversion comprising of pseudogene harbouring mutations. The missense mutations (p.G19R, p.K355R, p.D437Y, p.C633R, p.M771V, p.G2044D, p.C2491R) appear to play a major role and were identified in seven (36.8%) patients. In conclusion, a high frequency of novel mutations suggests the high propensity of VWF for new mutations. Missense and deletion mutations found to be a common cause of VWD3 in cohort of Indian VWD3 patients. Breakpoints characterisation of two large deletions reveals the double strand break and non-homologous recombination as deletions mechanism.
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http://dx.doi.org/10.1160/TH12-10-0737DOI Listing
April 2013

Characterisation of mutations and molecular studies of type 2 von Willebrand disease.

Thromb Haemost 2013 Jan 22;109(1):39-46. Epub 2012 Nov 22.

Department of Hematology, All India Institute of Medical Sciences AIIMS, New Delhi 110029, India.

Type 2 von Willebrand disease (VWD) is characterised by qualitative defects in von Willebrand factor (VWF). Exon 28 of the VWF gene is known to be a hot spot for type 2 VWD mutations. The goal of this study was to characterise the mutations in VWF exon 28 and understand the molecular basis of phenotypes through in vitro and in silico studies. Mutation screening was performed in 56 type 2 VWD patients through direct sequencing. Expression vectors for five mutations were transiently expressed in 293-EBNA cells to understand the mutations pathology. Furthermore, in silico structure analysis was performed for 13 missense mutations. A total of 16 including eight novel mutations were detected in 23 (41%) patients. Of these, 15 were missense (including seven V1439M, A1464P, M1495L, I1509V, R1527Q, N1635I and A1647D novel ones) and one was a novel gene conversion. Expression studies and characterisation of recombinant VWF suggested the loss of VWF function for mutants P1266Q, V1439M and N1635I and gain of function for mutant R1308C. No apparent defect was seen in mutant N1231S. In silico structure analysis suggested the probable gain or loss of hydrogen/van der Waals interactions in 10 mutant proteins. In conclusion, type 2A mutations and gene conversion were found to be a common cause of type 2 VWD. Expression studies suggest the mutations N1635I for type 2A(II), P1266Q and V1439M for type 2M, R1308C for type 2B VWD and N1231S as a non-causative variant. Moreover, in silico studies of the mutants show the probable cause of respective phenotypes.
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http://dx.doi.org/10.1160/TH12-07-0475DOI Listing
January 2013

Omic approaches to quality biomarkers for stored platelets: are we there yet?

Transfus Med Rev 2010 Jul;24(3):211-7

Section of Cell Biology, Division of Hematology, Office of Blood Research and Review, Center for Biologics Evaluation and Research, US Food and Drug Administration, Bethesda, MD, USA.

At present, there is no single biomarker that serves as the "gold standard" predictive of the quality of stored platelets used for transfusion. Some of the measurable features of platelets such as morphology, biochemical status, physiologic response to osmotic stress and agonist-induced changes, and measurement of process-associated activation indicators of platelets are considered useful in assessing the in vitro quality of stored platelets. Such in vitro measurements combined with in vivo survival estimations using radiolabeled platelets in healthy volunteers provide reasonable estimates of in vivo platelet function after transfusion. Thus, the current practice of estimating the quality and functional aspects of ex vivo stored platelets involves utilization of a battery of tests that dates back to pre-omic era. On the other hand, during the last decade, seminal discoveries have been made in platelet molecular and cell biology by using "omic"-based approaches such as proteomics, genomics, and transcriptomics. Can we mobilize some of these discoveries toward developing reliable quality biomarkers for stored platelets? To address this topic, we briefly review current practices and provide insights into some of the omic approaches that could be helpful in identifying quality storage biomarkers of platelets in the near future. We also briefly discuss here some of the challenges in using proteomic approaches and advantages of using one of the transcriptomics approaches toward platelet biomarker development.
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http://dx.doi.org/10.1016/j.tmrv.2010.03.003DOI Listing
July 2010

Differential profiling of human red blood cells during storage for 52 selected microRNAs.

Transfusion 2010 Jul 11;50(7):1581-8. Epub 2010 Feb 11.

Section of Cell Biology, Laboratory of Cellular Hematology, Center for Biologics Evaluation and Research, FDA, Bethesda, Maryland, USA.

Background: MicroRNAs (miRNAs), the negative regulators of cellular mRNAs, are present in mature red blood cells (RBCs) in abundance relative to other blood cells. So far, there are no studies aimed at identifying large-scale miRNA profiles during storage of RBCs.

Study Design And Methods: RNA samples from each RBC bag stored at 4 degrees C were collected on Days 0, 20, and 40 and subjected to miRNA profiling by using a membrane-based array. Fifty-two selected miRNAs of cellular apoptotic pathway represent the array. Through bioinformatics analyses, we identified potential target genes for selected miRNAs.

Results: Differential profiling of RBCs for 52 miRNAs revealed two distinguishable patterns during storage: Forty-eight miRNAs demonstrated no trend at all, while four miRNAs, miR-96, miR-150, miR-196a, and miR-197, demonstrated an increase up to Day 20 and subsequently decreased during storage. We selected miR-96 and subjected it to standard bioinformatics analyses for target gene predictions, which identified several mRNAs including the RBC proapoptotic calpain small subunit-1 (CAPNS1) as potential targets of miR-96. To validate these predictions, we selected CAPNS1 mRNA as an example and confirmed its presence in the RBCs. Future experimental verification would help define miR-96-CAPNS1 interaction, if any, in the stored RBCs.

Conclusions: This study for the first time provided a differential profile of stored RBCs for selected miRNAs related to cellular apoptotic pathway and opened new avenues toward identification of novel in vitro RBC biomarkers of storage lesions. Future studies focusing on target gene-miRNA interactions in stored RBCs would also unravel underlying mechanisms of storage lesions.
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http://dx.doi.org/10.1111/j.1537-2995.2010.02585.xDOI Listing
July 2010

Coinheritance of severe von Willebrand disease with Glanzmann thrombasthenia.

Clin Appl Thromb Hemost 2010 Oct 10;16(5):529-32. Epub 2010 Feb 10.

Department of Hematology, IRCH Building, 1st Floor, All India Institute of Medical Sciences (AMS), New Delhi, India.

A 35-years old male patient presented severe bleeding was diagnosed to have type 3 von Willebrand disease (VWD) and carrier for Glanzmann thrombasthenia (GT). Propositus and family members were studied through basic coagulation tests and genomic DNA analysis. Two offspring of the family were diagnosed to have GT through platelet aggregation along with VWD carrier. The patient with VWD was found positive for homozygous truncating mutation R1659X in VWF gene, and all offspring were heterozygous carriers of null allele. Hence, propositus was a carrier of GT with severe type 3 VWD and wife was a carrier of GT. Thus, it is concluded that there is importance of careful studies of patients even from nonconsanguineous families to exclude unusual coinheritance of congenital hemostatic disorders. If single replacement therapy in patient not responding well then probably co-expression of coagulopathies required and multiple replacement therapy should be given according to clinical and laboratory features.
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http://dx.doi.org/10.1177/1076029609360527DOI Listing
October 2010

Impact of thrombogenic mutations on clinical phenotypes of von Willebrand disease.

Clin Appl Thromb Hemost 2010 Jun 2;16(3):281-7. Epub 2009 Dec 2.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

von Willebrand disease (VWD) is a most common inherited bleeding disorder. von Willebrand factor (VWF) exists as an extracellular adaptor molecule and generally involves in the hemostasis mechanism through binding with GP (Glycoprotein) Ib-IX-V platelet receptor. Clinical phenotype of bleeding disorders modulated to a decrease in bleeding symptoms by thrombogenic mutations. We made an attempt to investigate the impact of thrombogenic mutations/polymorphisms on the clinical phenotype of 114 different types of patients with VWD, and 120 healthy controls were screened for methylenetetrahydrofolate reductase (MTHFR) 677C/T, factor V (FV) Leiden (1691G/A), beta(3) integrin (HPA-I) (Human platelets antigen-I) gene (1565T/C), and prothrombin 20210G/A mutations. Genotypic analysis was performed using polymerase chain reaction (PCR) and restriction fragment length polymorphism. Forty-five patients (39.5%) were found to be positive for at least one of the prothrombotic risk factors screened. Prothrombin 20210G/A was not found in any patient with VWD as well as healthy control. Eight patients with VWD were carrying the defective alleles of different thrombogenic markers, showing milder phenotypes than expected. A high prevalence was observed for MTHFR 677C/T (677C/C 73.6%, 677C/T 24.6%, 677T/T 1.8%) and PLA1/A2 (1565T/T 88.6%, 1565T/C 10.5%, 1565C/C 0.87%) polymorphism followed by FV Leiden (1691G/G 97.4%, 1691G/ A 2.6%, 1691A/A 0.00%) in patients with VWD with allelic frequencies 11.4% (677T), 5% (1565C), and 1.3% (1691A). Hence, we concluded that thrombophilic markers were seen to be influencing the clinical phenotypes of patients with VWD.
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http://dx.doi.org/10.1177/1076029609351291DOI Listing
June 2010

An update on the prevalence and characterization of H-PF4 antibodies in Asian-Indian patients.

Semin Thromb Hemost 2009 Apr 18;35(3):337-43. Epub 2009 May 18.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Heparin is the second most widely used anticoagulant/antithrombotic agent besides warfarin and is commonly used for various purposes such as treatment and surgical indications. A significant adverse effect of heparin treatment can occur when heparin binds platelet factor 4 (H-PF4) to form a complex that results in formation of H-PF4 antibodies, which in turn leads to platelet/endothelial cell activation followed by heparin-induced thrombocytopenia (HIT). Based on the heparin-induced platelet aggregation and enzyme-linked immunosorbent assay tests, the H-PF4 antibody (HIT antibody) was diagnosed in 6% of Indian patients undergoing cardiovascular surgery who received unfractionated heparin, but the frequency of occurrence rose to 15% when the patients were tested with the (14)C-serotonin release assay. This highlights some methodological variations in the diagnosis of HIT antibodies. It was also found that all the HIT-positive patients were either homozygous or heterozygous for the FcgammaRIIa polymorphism, which also highlights the role of this polymorphism in the occurrence of HIT. Very recent studies show that increases in HIT antibody production may also be due to heparin contaminants. Although these antibodies do not result in thrombocytopenia, contaminants in heparin may be capable of triggering a differential immunogenic response in comparison with contaminant-free heparin. Here we discuss the methodological differences in diagnosing HIT, the potential impact of contaminants in heparin, as well as future considerations.
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http://dx.doi.org/10.1055/s-0029-1222612DOI Listing
April 2009

Membrane array-based differential profiling of platelets during storage for 52 miRNAs associated with apoptosis.

Transfusion 2009 Jul 31;49(7):1443-50. Epub 2009 Mar 31.

Section of Cell Biology, Laboratory of Cellular Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration/PHS, Bethesda, Maryland.

Background: Enucleated platelets (PLTs) utilize posttranscriptional gene (mRNA) regulation (PTGR) for their normal morphologic and physiologic functions, which are altered in their ex vivo storage, also collectively referred to as storage lesions. While cellular micro-RNAs (miRNAs) play a significant role in posttranscriptional gene (mRNA) regulation by binding to their target mRNAs, comprehensive analysis of apoptosis-associated miRNAs and global changes in their profiles during PLT storage have not been evaluated to date.

Study Design And Methods: In this report room temperature-stored PLTs of Days 0, 2, and 9 were analyzed by differential profiling for 52 apoptosis-associated human miRNAs. After total RNA extraction from the samples, a membrane array-based miRNA analysis was carried out. Prediction of target genes was performed by bioinformatics-based approaches.

Results: Our analysis revealed that during storage, Let-7a, -7c, -7e, -7f, -7g, and -7i miRNA profiles of the PLTs were barely detectable, while levels of miR-150, -151, -152, -184, -188, -196a, -197, and -202 remained at high levels in PLTs. The rest of the miRNA levels were in between. However, two miRNAs, Let-7b and miR-16, distinctly demonstrated an increasing trend while miR-7 and miR-145 showed a decreasing profile during PLT storage. For these four miRNAs, we also identified their potential target mRNAs.

Conclusions: Overall, these results confirm the fact that miRNAs do exist in PLTs, and among 52 apoptosis-specific miRNAs studied, only a few selected miRNAs did perturb during PLT storage. Future experimental evaluation of these miRNA-target mRNA interactions will provide new insights into the molecular mechanisms of PLT storage-associated lesions.
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http://dx.doi.org/10.1111/j.1537-2995.2009.02140.xDOI Listing
July 2009

Increased prevalence of antiheparin platelet factor 4 antibodies in patients may be due to contaminated heparin.

Clin Appl Thromb Hemost 2009 Mar-Apr;15(2):145-51

Department of Pathology, Loyola University Medical Center, Maywood, Illinois 60153, USA.

During the period of November 2007 to January 2008, an increased prevalence of adverse reactions to heparin was noted. These adverse events have been attributed to the presence of purposeful contaminant, oversulfated chondroitin sulfate (OSCS) from April 2007 to May 2008. An analysis of dialysis patients' plasma obtained in 2006 and 2007 consistently had a low (5%) prevalence of AHPF4 antibodies. Blood samples from 78 patients on maintenance hemodialysis, who were potentially exposed to OSCS-contaminated heparin, were analyzed for the presence of all AHPF4 antibodies using a commercially available ELISA kit from GTI. Although there was no change in the platelet count of these patients, 15 of 78 patients (19.2%) studied had an increased prevalence of AHPF4 antibodies. Subtyping of the all platelet factor 4 (PF4) antibodies documented showed a higher prevalence of immunoglobulin G antibodies as compared to their previously determined antibodies. These observations suggest that the OSCS contaminant in the recalled heparin triggers an immunogenic response not seen with OSCS-contaminated free heparin.
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http://dx.doi.org/10.1177/1076029609332802DOI Listing
September 2009

Modulation of clinical phenotype of Glanzmann's thrombasthenia by thrombogenic mutations.

Clin Chim Acta 2009 May 24;403(1-2):156-8. Epub 2009 Feb 24.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Background: Glanzmann's thrombasthenia (GT) is an autosomal recessive bleeding disorder which is due to a defect in platelet aggregation in response to multiple physiological agonists. It has been demonstrated that the clinical phenotype of various diseases inherited in a classic Mendelian fashion can be modulated by a series of factors, inherited as well as acquired.

Methods: A total of 45 GT patients were screened for the thrombogenic polymorphisms, i.e., FV Leiden (R506Q), Prothrombin G20210A, MTHFR C677T and HPA-1 by PCR/RFLP.

Results: MTHFR C677T heterozygous was seen in 6 patients, FV Leiden heterozygous in one and Prothrombin G20210A gene variant in none. HPA-1 was seen in 3 patients of whom 1 was homozygous and 2 were heterozygous.

Conclusion: Thus the coinheritance of heterozygous FV Leiden alone or homozygous HPA 1b alone or the combined heterozygosity of MTHFR and HPA-1 were predicted to alter the clinical phenotype. Whereas the inheritance of heterozygous MTHFR alone or heterozygous HPA-1 alone did not altered the clinical phenotype significantly. Hence FV Leiden, MTHFR C677T polymorphism along with PLA-1 and homozygous HPA-1 were the probable ameliorating factor in GT mild phenotype.
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http://dx.doi.org/10.1016/j.cca.2009.02.009DOI Listing
May 2009

Glanzmann's thrombasthenia in North Indians: sub classification and carrier detection by flow cytometry.

Platelets 2009 Feb;20(1):12-5

Department of Haematology, All India Institute of Medical Sciences, New Delhi, India.

Thirty-three patients of Glanzmann's thrombasthenia (GT) and their families were assessed for the expression of alphaIIbbeta3 on platelet surface, by flow cytometry, to determine the common subtypes in North Indians as well as to assess the carrier status in family members of GT patients. GT was diagnosed in patients with bleeding manifestations accompanied by absent/reduced platelet aggregation, secondary to adenosine-di-phosphate, adrenaline, arachidonic acid and collagen. Based on alphaIIbbeta3 levels, 21 patients (64%) were classified as type I (as alphaIIbbeta3 was absent), 4 patients (12%) as type II and 8 patients (24%) as type III. Eight out of 20 fathers, 10 out of 20 mothers and 20 out of 31 siblings were found to have reduced alphaIIbbeta3 levels. Reduced alphaIIbbeta3 expression was seen in 63% of parents and 65% of siblings. It is possible that low alphaIIbbeta3 levels in family members may reflect their carrier status. It is postulated that flow cytometry estimation of alphaIIbbeta3 in parents/siblings may detect carrier status in GT. It is also revealed that type I GT is the commonest subtype in North Indians.
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http://dx.doi.org/10.1080/09537100802434853DOI Listing
February 2009

Glanzmann's thrombasthenia: an overview.

Clin Appl Thromb Hemost 2009 Mar-Apr;15(2):152-65. Epub 2008 Oct 16.

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Glanzmann's thrombasthenia (GT) is an autosomal recessive inherited bleeding disorder due to a defect in platelet function. The hallmark of this disease is severely reduced/absent platelet aggregation in response to multiple physiological agonists. Bleeding signs in GT include epistaxis, bruising, gingival hemorrhage, gastrointestinal hemorrhage, hematuria, menorrhagia, and hemarthrosis. Homozygous or compound heterozygous mutations in the genes of GPIIb and GPIIIa lead to GT. A patient with GT, with no possible causative mutations in GPIIb and GPIIIa genes, may harbor defects in a regulatory element affecting the transcription of these 2 genes. GT occurs in high frequency in certain ethnic populations with an increased incidence of consanguinity such as in Indians, Iranians, Iraqi Jews, Palestinian and Jordanian Arabs, and French Gypsies. Carrier detection in GT is important to control the disorder in family members. Carrier detection can be done both by protein analysis and direct gene analysis.
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http://dx.doi.org/10.1177/1076029608326165DOI Listing
September 2009

Impact of 789Ala/Ala genotype on quantitative type of von Willebrand disease.

Ann Hematol 2009 May 16;88(5):479-83. Epub 2008 Oct 16.

Department of Haematology, All India Institute of Medical Sciences, New Delhi, India.

von Willebrand factor (VWF) is a complex multimeric plasma glycoprotein encoded by an approximately 178-kb large VWF gene located on the short arm of chromosome 12 (12p13.2). VWF plays an important role in hemostasis through binding with platelet GpIbalpha receptors. We made an attempt to correlate the 789Ala/Ala genotype of the VWF with VWF:Ag level in different types of unrelated von Willebrand disease (VWD) patients and healthy controls. VWF assays and other coagulation screening tests have been done for all 103 (50 male, 53 female) different types of index VWD patients including 19 type 1, 55 type 2, and 29 type 3 VWD patients. Genotypes were detected by polymerase chain reactions followed by restriction fragment length polymorphism. The genotype 789Ala/Ala was found in 26.3% type 1 and in 31.0% type 3 patients. This genotype was not found in any of type 2 patient or healthy controls. Overall, 789Ala/Ala genotype was found significantly higher (P < 0.001) in quantitative type (type 1 and type 3) VWD that is occurred due to low VWF:Ag level. These results demonstrate that mutant homozygous 789Ala/Ala genotype of this polymorphism probably have their functional implications for low plasma VWF:Ag level in quantitative type of VWD.
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http://dx.doi.org/10.1007/s00277-008-0623-4DOI Listing
May 2009

Carrier detection in Glanzmann thrombasthenia: comparison of flow cytometry and Western blot with respect to DNA mutation.

Am J Clin Pathol 2008 Jul;130(1):93-8

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

We studied 20 families with Glanzmann thrombasthenia, including 20 parents and 22 siblings, for carrier detection. Carrier detection was done by phenotypic analysis (flow cytometry and Western blot) and direct gene analysis (sequencing or restriction fragment length polymorphism). Reduced expression of the glycoprotein IIb/IIIa complex was found by flow cytometry in 17 (85%) of the parents and 12 (55%) of the siblings. Western blot showed a reduced or an abnormal band in 6 (30%) of the parents and 8 (36%) of the siblings. DNA analysis in family members showed all parents and 16 (73%) of siblings to be carriers. Both techniques, flow cytometry and Western blot, were evaluated with respect to DNA analysis as a "gold standard" method. The sensitivity of flow cytometry was higher (75%) than that of Western blot (39%). We concluded that flow cytometry can effectively be used for carrier detection in Glanzmann thrombasthenia.
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http://dx.doi.org/10.1309/HYE4AP9961CEP0C0DOI Listing
July 2008

Hypercoagulable state in five thalassemia intermedia patients.

Clin Appl Thromb Hemost 2007 Oct;13(4):422-7

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Fifty-three patients of thalassemia intermedia and 40 controls were studied for clinical evidence of thrombosis and laboratory evidence of hypercoagulable state. Thrombotic episodes were detected in 5 (9.4%) patients. Two of these 5 patients with thrombosis were splenectomized. Laboratory evaluation showed presence of thrombocytosis in 8 (15%), 5 of these were splenectomized. Platelet hyperaggregation was detected in 12 (22.2%) patients. Although rate of aggregation was slow in 7 (13.2%) patients, degree of aggregation was normal in these 7 patients and platelet hypoaggregation was not detected in any patient. Level of coagulation inhibitors protein C and protein S, and antithrombin III were decreased in 31 (58.4%) patients. There was no correlation between low level of protein C and protein S with hepatic dysfunction and iron overload. Antithrombin III level was decreased only in 8 (15%) patients. There was a statistically significant association between the lower level of this inhibitor and hepatic dysfunction. In conclusion, this study provides evidence for the existence of a chronic hypercoagulable state in patients with beta thalassemia intermedia, and suggests that expression of a procoagulant surface by thalassemia intermedia red blood cells may be the major underlying factor giving rise to platelet and coagulation inhibitor abnormalities in these patients. These alterations are not related to iron overload or hepatic dysfunction.
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http://dx.doi.org/10.1177/1076029607303539DOI Listing
October 2007

Inherited platelet function disorders versus other inherited bleeding disorders: an Indian overview.

Thromb Res 2008 11;121(6):835-41. Epub 2007 Sep 11.

Department of Haematology, IRCH Building 1st floor, All India Institute of Medical Sciences, Ansari Nagar, New Delhi-110 029, India.

Inherited deficiencies of plasma proteins involved in blood coagulation generally lead to lifelong bleeding disorders, whose severity is directly proportional to the degree of factor deficiency. Platelet and other coagulation factors play an important role in the haemostasis mechanism. We attempted to study the prevalence of inherited platelet function disorders (PFDs) and correlate with other coagulation factor disorders in the Indian population. Patients with PFDs and other coagulation factor disorders who presented at our hospital during the 5 year period (from January, 2001 to December, 2005) were the subjects of the study. A total of 1576 patients were diagnosed to have congenital bleeding disorders including PFDs, von Willebrand disease, haemophilia A and B and rare coagulation disorder cases. Haemophilia A (HA) was the most common and was seen in 52.31% of the patients followed by total PFDs seen in 27.77% of the patients. Based on severity of the disease, the results of PFDs were highly significant when compared to haemophilia and von Willebrand disease (VWD) (p=0.000). Severity was found higher in HA (77.8%) followed by HB (69.6%) and was found lower for PF3 availability defect (9.0%). It has been concluded that the prevalence of PFDs is relatively low as compared to coagulation factors related disorder and also it has been established that type-1 VWD is relatively less frequent in India as compared to the West.
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http://dx.doi.org/10.1016/j.thromres.2007.07.015DOI Listing
October 2008

Laboratory studies in coagulation disorders.

Indian J Pediatr 2007 Jul;74(7):649-55

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

It is important to go in a stepwise approach to diagnose spectrum of bleeding disorders, so that minimum tests are undertaken to make a definitive diagnosis and to avoid unnecessary tests and laboratory load. Depending on the abnormalities observed in the short screening, extended screening tests can be performed followed by specialized diagnostic tests. Bleeding time is prolonged in thrombocytopenia and platelet function disorders (PFD). If platelet count is normal, extended screening tests such as RVVT, PF3 availability and clot retraction can be performed. Russel viper venom directly activates FX, in presence of PF3, is an indicator of common pathway of coagulation. However, if there is deficiency of PF3 as obtained in PFD and APTT PT are normal, its prolongation indicates PFD. These can be tested invitro by performing RVVT with and without inosithin it is highly suggestive of underlying PFD. In such cases, diagnostic tests for PFD such as platelet aggregation with ADP, ADR, AA, Collagen and Ristocetin can be performed followed by electron microscopy if possible. Few of the interesting cases also have been discussed in the text.
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http://dx.doi.org/10.1007/s12098-007-0116-9DOI Listing
July 2007

Disseminated intravascular coagulation in acute leukemia at presentation and during induction therapy.

Clin Appl Thromb Hemost 2007 Jul;13(3):292-8

Department of Haematology, All India Institute of Medical Sciences, New Delhi, India.

Between January 2001 and December 2003, 67 patients with acute leukemia were evaluated prospectively for hemostatic abnormality at presentation, of which 43 (64.2%) had acute lymphoblastic leukemia and 24 (35.8%) had acute myelogenous leukemia. At presentation, 27 patients (40.3%) had bleeding manifestations. Thrombocytopenia was present in 57 patients (85%), and 33(49.3%) had some abnormality of global coagulation markers. Disseminated intravascular coagulation was defined by International Society of Thrombosis and Hemostasis criteria. Disseminated intravascular coagulation was more often associated with bleeding manifestations in acute myelogenous leukemia cases than in acute lymphoblastic leukemia cases. Two patients presented disseminated intravascular coagulation on day 7 of chemotherapy, without any bleeding manifestations. Four of 15 evaluated cases who had a bleeding or infection complication after day 7 of induction therapy also had disseminated intravascular coagulation. It is recommended that all patients with leukemia be investigated for disseminated intravascular coagulation at presentation.
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http://dx.doi.org/10.1177/1076029607302435DOI Listing
July 2007

Therapy-related acute promyelocytic leukemia after treatment of carcinoma breast--a case report.

Indian J Pathol Microbiol 2006 Apr;49(2):251-4

Department of Haematology, All India Institute of Medical Sciences, New Delhi.

We report a 43-year-old female, with acute promyelocytic leukemia occurring after 9 months of treatment for carcinoma breast. The diagnosis of APL was made on morphology, cytogenetics and molecular studies. In contrast to other published report of therapy related APL (tAPL) the present case presented early after the primary malignancy and underwent a rapid, downhill course.
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April 2006

Gel card in the diagnosis of autoimmune haemolytic anemia.

Indian J Pathol Microbiol 2005 Jul;48(3):322-4

Department of Haematology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi.

The diagnosis of autoimmune haemolytic anaemia (AIHA) requires the establishment of haemolysis and demonstration of autoantibodies against red cells. Most laboratories use the conventional Coomb's test for the demonstration of the autoantibodies. However, in approximately 2-6% of the patients who present with the clinical and haematological features of AIHA, the direct agglutination test is negative on repeated testing. Attempts are therefore being made to identify a test which could be more sensitive than the conventional test, yet retaining the simplicity and cost effectiveness of the test. In the present study, the efficacy of the newly developed gel card test has been compared with the conventional Coomb's test for detection of autoantibodies in 50 cases clinically suspected to have haemolytic anemia. The gel card picked up the antibodies in all the cases detected to be positive by the conventional test. In addition, the gel card also picked up 5 tests which were negative by the conventional method. The sensitivity and specificity of the gel card Direct Coomb's test (DCT) as compared to the conventional tube test for DCT was found to be 100% and 95.1% respectively. The Indirect Coomb's test (ICT) was 100% sensitive and 92.5% specific. In view of the high sensitivity and specificity and the simplicity of the procedure, this test may be effectively used for diagnosis of AIHA.
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July 2005

Roles of protein C, protein S, and antithrombin III in acute leukemia.

Am J Hematol 2006 Mar;81(3):171-4

Department of Hematology, All India Institute of Medical Sciences, New Delhi, India.

Protein C, protein S, and antithrombin III were measured in 35 patients with acute leukemia (13 with AML and 22 with ALL). Low levels of proteins C and S were present in 15 (42.9%) and 20 (57.1%) patients, respectively, and 6 patients had low levels of antithrombin (ATIII). Seven patients also had DIC at presentation. There were no significant differences in the levels of protein C, protein S, and ATIII in patients with or without DIC. Twenty patients were available for re-evaluation at the end of induction therapy. The low levels of protein C and ATIII found at diagnosis had risen to normal levels at the end of the induction therapy, while low =levels of protein S remained in 75% of the patients. One patient with low protein C at presentation developed myocardial infarction on day 15, and another patient died of progressive neuropathy. No other thrombotic manifestations were seen. Whether the low protein C, protein S, or antithrombin levels predispose patients with acute leukemia to thrombosis in the absence of DIC is not known.
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http://dx.doi.org/10.1002/ajh.20546DOI Listing
March 2006
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