Predicting 2-Day Mortality of Thrombocytopenic Patients Based on Clinical Laboratory Data Using Machine Learning.

BACKGROUND: Clinical laboratories have traditionally used a single critical value for thrombocytopenic events. This system, however, could lead to inaccuracies and inefficiencies, causing alarm fatigue and compromised patient safety. OBJECTIVES: This study shows how machine learning (ML) models can provide auxiliary information for more accurate identification of critical thrombocytopenic patients when compared with the traditional notification system. RESEARCH DESIGN: A total of 50,505 patients' platelet count and other 26 additional laboratory datasets of each thrombocytopenic event were used to build prediction models. Conventional logistic regression and ML methods, including random forest (RF), artificial neural network, stochastic gradient descent (SGD), naive Bayes, support vector machine, and decision tree, were applied to build different models and evaluated. RESULTS: Models using logistic regression [area under the curve (AUC)=0.842], RF (AUC=0.859), artificial neural network (AUC=0.867), or SGD (AUC=0.826) achieved the desired average AUC>0.80. The highest positive predictive value was obtained by the SGD model in the testing data (72.2%), whereas overall, the RF model showed higher sensitivity and total positive predictions in both the training and testing data and outperformed other models. The positive 2-day mortality predictive rate of RF methods is as high as 46.1%-significantly higher than using the traditional notification system at only 14.8% [χ2(1)=81.66, P<0.001]. CONCLUSIONS: This study demonstrates a data-driven ML approach showing a significantly more accurate 2-day mortality prediction after a critical thrombocytopenic event, which can reinforce the accuracy of the traditional notification system.

Identification of a novel A2 allele through nt543 substitution.

BACKGROUND: ABO blood system has many subgroups. In A group, A1 phenotype and A2 phenotype are more common, and A2 is caused by deletion or substitution in A1 allele (ABO*A1.01). METHODS: Based on standard ABO serological test, the subject was identified as A2 phenotype. Direct sequencing and ABO gene cloning were performed to analyze the allele. RESULTS: The subject had one A1v allele (ABO*A1.02) and one O allele. The haplotype sequencing analysis of each allelic clone demonstrated that allele 1 was A1v (ABO*A1.02) allele with nt543 variation (543 G > C) and allele 2 was O1v allele (ABO*O.01.02) with nt261 deletion and nt220 variation. CONCLUSION: The 543 G > C nucleotide substitution of the present A1v allele (ABO*A1.02) shares the same sequence variation site with Ax allele (ABO*AW.33) (543 G > T), and both 543 G > C and 543 G > T nucleotide substitutions encode the same amino acid change of tryptophan to cysteine. Mechanism, such as allelic enhancement, has been proposed to explain this controversial phenotype-genotype relationship. But in present case, there has been no B allele to enhance the expression of Ax to that expected of A2, so there could be another novel underlying mechanism to be investigated.

Rapid rare ABO blood typing using a single PCR based on a multiplex SNaPshot reaction.

BACKGROUND: ABO subgroups would be considered when discrepancies in ABO grouping occur. Serological methods including adsorption-elution test, salivary ABH inhibition test, and anti-A1 (lectin) saline method could be used. However, these serological methods are laboring and obscure. Therefore, reliable and affordable method to assess the ABO subgroups is of particular interest. METHODS: To solve this problem, the multiplex SNaPshot-based assays were designed to determine rare A and B subgroups. Primers used as probes for determination of rare ABO blood groups known in Taiwanese population were designed. Many ABO subtype samples were used to validate the accuracy and reproducibility of our SNaPshot panel. RESULTS: A panel of primer probes were successfully designed in determining 8 SNP sites (261, 539, 838, 820, 745, 664, IVS6 +5, and 829 in exon 6 and 7) for A phenotype and 6 SNP sites (261, 796, IVS3 +5, 247, 523, and 502 in exon 2, 6 and 7 and intron 3) for B phenotype. SNaPshot analysis for defining blood group A alleles (A1, A2, A3, Am and Ael) and blood group B alleles (B1, B3, Bw and Bel) was therefore available. CONCLUSION: SNaPshot analysis could be used in reference laboratories for typing known rare subgroups of A and B without DNA cloning and traditional sequencing. Moreover, this method would help to construct databases of genotyped blood donors, and it potentially plays a role in determining fetal-maternal ABO incompatibility.

Association between diminished miRNA expression and the disease status of AML patients: Comparing to healthy control.

BACKGROUND: Changes in ABO blood type caused by a gradual decrease in antigen expression have been found in patients with acute myeloid leukemia (AML). Studies have indicated that alteration of ABO gene methylation accounts for 50% of acquired weak ABO antigen expression in patients with leukemia. However, the molecular mechanisms contributing to the remaining 50% of cases are unknown. We hypothesize that deregulation of miRNA is correlated with weak ABO antigen expression in patients with AML. METHODS: Blood samples of 19 patients with AML and 12 healthy controls were collected, in which the blood type was not changed in these AML patients. Flow cytometric analysis was applied to measure the ABO antigen expression titer among AML patients and controls. A total of 18 leukemia-related miRNAs were analyzed via quantitative real-time polymerase chain reactions. RESULTS: We found that miRNA profiles were correlated with the AML patients, especially in those who had constant or weakened ABO antigen expressions. Compared with healthy controls, the miR-16 and miR-451 expression were significantly lower in either AML cases with weak ABO antigen expressions (p = 0.003, p = 0.028, respectively) or AML cases with constant ABO antigen expressions (p = 0.043, p = 0.040, respectively). Although not statistically significant, decreasing trends in the miR-451 and miR-16 expressions in the AML patients with weakened ABO were observed compared to those with constant ABO antigens. The weak ABO antigen expression might correlate with miRNAs, especially miR-16 and miR-451. CONCLUSION: This study indicated that decreasing in miR-16 and miR-451 was associated with AML and AML with weakened ABO expression. In the future, we will continue to include more cases and exclude the others factor influencing ABO antigen expression, promoter methylation and oxidative stress, to replicate the results of this study and investigate the underlying mechanism of decreasing miR-16 and miR-451 in AML patients with varied ABO antigen expression levels.

Exploring the Bio-Functional Effect of Single Nucleotide Polymorphisms in the Promoter Region of the TNFSF4, CD28, and PDCD1 Genes.

In a prior study, we discovered that hematopoietic stem cell transplantation (HSCT) and/or autoimmune diseases, such as systemic lupus erythematosus, were associated with the rs1234314 C/G and rs45454293 C/T polymorphisms of TNFSF4, the rs5839828 C > del and rs36084323 C > T polymorphisms of PDCD1, and the rs28541784C/T, rs200353921A/T, rs3181096C/T, and rs3181098 G/A polymorphisms of CD28. However, the association does not imply causation. These single nucleotide polymorphisms (SNPs) are all located in the promoter region of these genes, so we used the dual-luminescence reporter assay to explore the effect of single nucleotide polymorphisms (SNPs) on transcriptional activity. For each promoter-reporter with a single SNP mutation, more than 10 independent experiments were carried out, and the difference in transcription activity was compared using one-way ANOVA and Tukey's honestly significant difference test. The results showed that the G-allele of rs1234314 had 0.32 ± 0.09 times the average amount of relative light units (RLU) compared to the C-allele (p = 0.003), the T-allele of rs45454293 had 4.63 ± 0.92 times the average amount of RLU compared to the C-allele (p < 0.001), the del-allele of rs5839828 had 1.37 ± 0.24 times the average amount of RLU compared to the G-allele (p < 0.001), and the T-allele of rs36084323 had 0.68 ± 0.07 times the average amount of RLU compared to the C-allele (p < 0.001). The CD28 SNPs studied here did not affect transcriptional activity. In conclusion, the findings of this study could only confirm that the SNP had a bio-functional effect on gene expression levels. According to the findings, several SNPs in the same gene have bio-functions that affect transcriptional activity. However, some increase transcriptional activity while others decrease it. Consequently, we inferred that the final protein level should be the integration result of the co-regulation of all the SNPs with the effect on transcriptional activity.

Exploration of the association between the single-nucleotide polymorphism of co-stimulatory system and rheumatoid arthritis.

Introduction: The human leukocyte antigen (HLA) has been linked to the majority of autoimmune diseases (ADs). However, non-HLA genes may be risk factors for ADs. A number of genes encoding proteins involved in regulating T-cell and B-cell function have been identified as rheumatoid arthritis (RA) susceptibility genes. Methods: In this study, we investigated the association between RA and single-nucleotide polymorphisms (SNPs) of co-stimulatory or co-inhibitory molecules in 124 RA cases and 100 healthy controls without immune-related diseases [including tumor necrosis factor superfamily member 4 (TNFSF4), CD28, cytotoxic T-lymphocyte-associated protein 4 (CTLA4), and programmed cell death protein 1 (PDCD1)]. Results: The results showed that there were 13 SNPs associated with RA, including rs181758110 of TNFSF4 (CC vs. CT, p = 0.038); rs3181096 of CD28 (TT vs. CC + CT, p = 0.035; CC vs. TT, p = 0.047); rs11571315 (TT vs. CT, p = 0.045), rs733618 (CC vs. TT + CT, p = 0.043), rs4553808 (AA vs. AG vs. GG, p = 0.035), rs11571316 (GG vs. AG vs. AA, p = 0.048; GG vs. AG + AA, p = 0.026; GG vs. AG, p = 0.014), rs16840252 (CC vs. CT vs. TT, p = 0.007; CC vs. CT, p = 0.011), rs5742909 (CC vs. CT vs. TT, p = 0.040), and rs11571319 of CTLA4 (GG vs. AG vs. AA, p < 0.001; GG vs. AG + AA, p = 0.048; AA vs. GG + AG, p = 0.001; GG vs. AA, p = 0.008; GG vs. AG, p ≤ 0.001); and rs10204525 (TT vs. CT + CC, p = 0.024; TT vs. CT, p = 0.021), rs2227982 (AA vs. GG, p = 0.047), rs36084323 (TT vs. CT vs. CC, p = 0.022; TT vs. CT + CC, p = 0.013; CC vs. TT + CT, p = 0.048; TT vs. CC, p = 0.008), and rs5839828 of PDCD1 (DEL vs. DEL/G vs. GG, p = 0.014; DEL vs. DEL/G + GG, p = 0.014; GG vs. DEL + DEL/G, p = 0.025; DEL vs. GG, p = 0.007). Discussion: Consequently, these SNPs may play an important role in immune regulation, and further research into the role of these SNPs of immune regulatory genes in the pathogenesis of RA is required.

Screening of single nucleotide polymorphisms within HLA region related to hematopoietic stem cell transplantation using MassARRAY technology.

A growing number of studies showed that single nucleotide polymorphisms (SNPs) in the human leukocyte antigen (HLA)-related genes were associated with the outcome of hematopoietic stem cell transplantation (HSCT). Thus, other SNPs located nearby the classical HLA genes must be considered in HSCT. We evaluated the clinical feasibility of MassARRAY by comparing to Sanger sequencing. The PCR amplicons with each one of the 17 loci that were related to the outcomes of HSCT published by our previous study were transferred onto a SpectroCHIP Array for genotyping by mass spectrometry. The sensitivity of MassARRAY was 97.9% (614/627) and the specificity was 100% (1281/1281), where the positive predictive value (PPV) was 100% (614/614) and the negative predictive value (NPV) was 99.0% (1281/1294). MassARRAY is high-throughput, which can accurately analyze multiple SNPs at the same time. Based on these properties, we proposed that it could be an efficient method to match the genotype between the graft and the recipient before transplantation.

Investigation of the correlation between immune thrombocytopenia and T cell activity-regulated gene polymorphism using functional study.

Thrombocytopenia is a condition where the platelet count is under 100 × 109/L, which is caused by various disorders. However, the mechanism of thrombocytopenia is still unclear. Hence, we tried to investigate the correlation between immune thrombocytopenia (ITP) and single nucleotide polymorphisms (SNPs) of genes related to T cell activation. There were 32 ITP patients and 30 healthy controls enrolled in this study. PCR and sequencing were used to find out the significant SNPs, which we focused on the promoter region of CTLA4 and CD28. In this study, the ITP cases were divided into primary ITP group, secondary ITP group, and the combination of the two to the follow-up analysis. Moreover, dual-luciferase reporter assay was used to evaluate the transcription activity of the significant SNP. We found the - 1765_rs11571315 of CTLA4 gene was associated with primary ITP (p = 0.006), secondary ITP (p = 0.008), and the combination of the two (p = 0.003). Moreover, the -318_rs5742909 also had statistical significance in secondary ITP group that was only caused by autoimmune disease (p = 0.019). In functional study, the rs5742909 would decrease 19% of the transcription activity when it carried a T-allele at this position (p = 0.040). It was noted that CTLA4 gene polymorphism was related to ITP but not CD28. According to our results, we surmised that CTLA4 is involved in the pathogenesis of ITP, and the secondary ITP result from the lower CTLA4 expression that leads to T cell over-activation.

Risks in Induction of Platelet Aggregation and Enhanced Blood Clot Formation in Platelet Lysate Therapy: A Pilot Study.

Platelet concentrates (PCs) are widely used in regenerative medicine; as it is produced from freeze-thawing PC, platelet lysate (PL) has a longer shelf life. The thrombotic risk of PL therapy needs to be explored since PL and PC contain cytokines that contribute to platelet aggregation and thrombus formation. Whole blood samples of 20 healthy subjects were collected; PL was produced from PCs with expired shelf life through freeze-thawing. The direct mixing of PL with platelet-rich plasma (PRP) or whole blood was performed. In addition, rotational thromboelastometry (ROTEM) was used to investigate whether PL enhanced coagulation in vitro; the effects of fibrinogen depletion and anticoagulants were evaluated to prevent hypercoagulation. The results showed that PL induced platelet aggregation in both PRP and whole blood. In ROTEM assays, PL was shown to cause a significantly lower clotting onset time (COT) and clot formation time (CFT), and a significantly greater α angle and maximum clot firmness (MCF). Compared with the controls, which were 1:1 mixtures of normal saline and whole blood, fibrinogen depletion of PL showed no significant difference in CFT, α angle and MCF. Moreover, heparin- and rivaroxaban-added PL groups demonstrated no clot formation in ROTEM assays. Platelet lysate-induced hypercoagulability was demonstrated in vitro in the present study, which could be prevented by fibrinogen depletion or the addition of an anticoagulant.

Association between the side effect induced by COVID-19 vaccines and the immune regulatory gene polymorphism.

People often worry about the side effects after vaccination, reducing the willingness to vaccinate. Thus, we tried to find out the risk of single nucleotide polymorphism (SNP) vaccines to improve the willingness and confidence in vaccination. Allergic and inflammatory reactions are the common vaccine side effects caused by immune system overreaction. In addition, a previous study showed significantly higher frequency of febrile reactions to measles vaccines in American Indians than in Caucasian children, indicating that the side effects varied in accordance with genetic polymorphisms in individuals. Thus, SNPs of immune regulatory genes, cytotoxic T-lymphocyte-associated protein 4 (CTLA4), CD28, tumor necrosis factor ligand superfamily member 4 (TNFSF4) and programmed cell death protein 1 (PDCD1) were included in this study to analyze their association with vaccine side effects. Moreover, 61 healthy participants were asked on the number of doses they received, the brand of the vaccine, and the side effects they suffered. We found that several SNPs were associated with side effects after the first or second dose of mRNA or adenoviral vector vaccines. Furthermore, these SNPs were associated with several autoimmune diseases and cancer types; thus, they played an important role in immune regulation. Moreover, rs3181096 and rs3181098 of CD28, rs733618 and rs3087243 of CTLA, and rs1234314 of TNFSF4 were associated with mild vaccine side effects induced by mRNA and adenoviral vector vaccines, which would play a potential role in vaccine-induced immune responses and may further lead to fatal side effects. These results could serve as a basis for investigating the mechanism of vaccine side effects. Furthermore, it was hoped that these results would address public concerns about the side effects of the COVID-19 vaccination. In clinical application, a rapid screening test can be performed to assess the risk of vaccine side effects before vaccination and provide immediate treatment.

CD28 Gene Polymorphisms in the Promoter Region Are Associated with Transfusion Reactions: A Functional Study.

Transfusion reactions are mainly induced by the interaction of an antigen and antibody. However, transfusion reactions still occur with the implementing of crossmatching and usage of pre-storage leukoreduced blood products. The roles of CD28 and CTLA4 gene polymorphisms in transfusion reaction have been shown, and subjects with certain single nucleotide polymorphisms (SNPs) of the CD28 or CTLA4 gene had a significantly higher risk of transfusion reactions. In total, 40 patients with transfusion reactions after receiving pre-storage leukoreduced blood products were enrolled in this study. We focused on the SNPs located in the CD28 promoter region (rs1879877, rs3181096, rs3181097, and rs3181098) to find out the significant SNP. A luciferase reporter assay was used to investigate the expression level of protein affected by promoter SNP variation. We found that the polymorphism of rs3181097 was associated with transfusion reactions (p = 0.003 in additive model and p = 0.015 in dominant model). Consequently, we investigated the biological function in the CD28 promoter polymorphisms (rs1879877 G > T, rs3181096 C > T, rs3181097 G > A, and rs3181098 G > A) by using dual-spectral luciferase reporter assay. The results showed that the ex-pression level of CD28 was decreased under the effect of rs3181097 with A-allele. This suggested that rs3181097 may regulate immune response through decreasing CD28 protein expression and then lead to development of transfusion reactions.

The adverse events of haematopoietic stem cell transplantation are associated with gene polymorphism within human leukocyte antigen region.

Adverse reactions may still occur in some patients after receiving haematopoietic stem cell transplantation (HSCT), even when choosing a human leukocyte antigen (HLA)-matched donor. The adverse reactions of transplantation include disease relapse, graft-versus-host disease (GVHD), mortality and CMV infection. However, only the relapse was discussed in our previous study. Therefore, in this study, we investigated the correlation between the gene polymorphisms within the HLA region and the adverse reactions of post-HSCT in patients with acute leukaemia (n = 176), where 72 patients were diagnosed with acute lymphocytic leukaemia (ALL) and 104 were acute myeloid leukaemia (AML). The candidate single nucleotide polymorphisms were divided into three models: donor, recipient, and donor-recipient pairs and the data of ALL and AML were analysed individually. Based on the results, we found 16 SNPs associated with the survival rates, the risk of CMV infection, or the grade of GVHD in either donor, recipient, or donor-recipient matching models. In the ALL group, the rs209132 of TRIM27 in the donor group was related to CMV infection (p = 0.021), the rs213210 of RING1 in the recipient group was associated with serious GVHD (p = 0.003), and the rs2227956 of HSPA1L in the recipient group correlated with CMV infection (p = 0.001). In the AML group, the rs3130048 of BAG6 in the donor-recipient pairs group was associated with serious GVHD (p = 0.048). Moreover, these SNPs were further associated with the duration time of survival after transplantation. These results could be applied to select the best donor in HSCT.

Evaluation the diagnostic accuracy of albuminuria detection in semi-quantitative urinalysis.

BACKGROUND: Taiwan has the highest end-stage renal disease prevalence in the world, and the costs on the maintenance of dialysis imposes a great financial burden on National Health Insurance. Routine urinalysis provides an opportunity for the early detection of microalbuminuria. We evaluated the accuracy of semi-quantitative chemical methods from Siemens Novus Pro12 dipstick for albumin-creatinine ratio (ACR). METHODS: We collected 1029 random urine samples and performed urinary analytic tests by Siemens Novus with Pro12 dipsticks and also calculated the urinary ACR. The reference method was performed by Hitachi LST008, a quantitative assay. The percentage of exact agreement in ACR was 81.9% between Siemens Novus and Hitachi LST008. The percentage of agreement within 1 level between the 2 methods was 98.5%. When ACR > 30 mg/g was defined as the threshold for positive results, the sensitivity, specificity, positive, and negative predictive values for microalbuminuria were 87.2%, 91.6%, 91.5%, and 87.3%, respectively. There were 778 cases with negative results of urinary protein, analyzed by conventional dipsticks. 149 of 778 (19.2%) cases were positive, measured by Pro12 dipsticks, and 111 of 149 (74.5%) cases were confirmed positive ACR by Hitachi LST008. CONCLUSIONS: Urinary ACR measured by Siemens Novus with Pro12 dipsticks was shown to be a reliable test for detection of microalbuminuria.

Diagnosis of Pneumocystis pneumonia by real-time PCR in patients with various underlying diseases.

BACKGROUND: Pneumocystis pneumonia (PCP) is a disease caused by the opportunistic infection of the fungus Pneumocystis jirovecii. Several PCR methods have been developed to aid in the diagnosis of PCP. In this study, we evaluated the performance of a real-time PCR in the diagnosis of PCP, in patients with various underlying diseases. METHODS: Ninety-seven BAL samples and 94 sputum samples from 191 patients were used in the study. Patients were classified as PCP (121 patients) or non-PCP (70 patients) based on their clinical and radiological presentations. RESULTS: Real time PCR amplified the P. jirovecii mitochondrial large-subunit rRNA gene with a detection limit of 68 copies of DNA per reaction. Non-PCP pathogens including 32 different fungi and bacteria were also evaluated. Overall, 71.9% of the samples from PCP patients and 14.5% of those from non-PCP patients were positive for the PCR test with a CT value of the real-time PCR below 45. The main underlying diseases of the patients were hematological or solid malignancies (47.1%) and HIV infection (8.9%). The CT values of the test were significantly lower in BAL samples from PCP patients than those from non-PCP patients (p = 0.024). No non-PCP patient had a CT value below 30, whereas samples from 24.8% of PCP patients with underlying diseases had a CT value below 30. CONCLUSION: Since false positive PCR results were obtained, perhaps due to colonization, we suggest that the diagnosis of PCP should be based on a combination of clinical symptoms, underlying diseases, and PCR results.

Improving Multi-Tumor Biomarker Health Check-up Tests with Machine Learning Algorithms.

BACKGROUND: Tumor markers are used to screen tens of millions of individuals worldwide at annual health check-ups, especially in East Asia. Machine learning (ML)-based algorithms that improve the diagnostic accuracy and clinical utility of these tests can have substantial impact leading to the early diagnosis of cancer. METHODS: ML-based algorithms, including a cancer screening algorithm and a secondary organ of origin algorithm, were developed and validated using a large real world dataset (RWD) from asymptomatic individuals undergoing routine cancer screening at a Taiwanese medical center between May 2001 and April 2015. External validation was performed using data from the same period from a separate medical center. The data set included tumor marker values, age, and gender from 27,938 individuals, including 342 subsequently confirmed cancer cases. RESULTS: Separate gender-specific cancer screening algorithms were developed. For men, a logistic regression-based algorithm outperformed single-marker and other ML-based algorithms, with a mean area under the receiver operating characteristic curve (AUROC) of 0.7654 in internal and 0.8736 in external cross validation. For women, a random forest-based algorithm attained a mean AUROC of 0.6665 in internal and 0.6938 in external cross validation. The median time to cancer diagnosis (TTD) in men was 451.5, 204.5, and 28 days for the mild, moderate, and high-risk groups, respectively; for women, the median TTD was 229, 132, and 125 days for the mild, moderate, and high-risk groups. A second algorithm was developed to predict the most likely affected organ systems for at-risk individuals. The algorithm yielded 0.8120 sensitivity and 0.6490 specificity for men, and 0.8170 sensitivity and 0.6750 specificity for women. CONCLUSIONS: ML-derived algorithms, trained and validated by using a RWD, can significantly improve tumor marker-based screening for multiple types of early stage cancers, suggest the tissue of origin, and provide guidance for patient follow-up.

Association of CTLA4 Gene Polymorphism with Transfusion Reaction after Infusion of Leukoreduced Blood Component.

Leukocytes and cytokines in blood units have been known to be involved in febrile non-hemolytic transfusion reaction (FNHTR), and these adverse reactions still occur while using pre-storage leukoreduced blood products. Blood transfusion is similar to transplantation because both implant allogeneic cells or organs into the recipient. CTLA4 gene polymorphism was found to be associated with graft-versus-host disease in hematopoietic stem cell transplantation. We performed a prospective cohort study at a major tertiary care center to investigate the correlation of CTLA4 gene polymorphism and transfusion reactions. Selected CTLA4 gene SNPs were genotyped and compared between patients with transfusion-associated adverse reactions (TAARs) and healthy controls. Nineteen patients and 20 healthy subjects were enrolled. There were 4 SNPs showing differences in allele frequency between patients and controls, and the frequency of "A" allele of rs4553808, "G" allele of rs62182595, "G" allele of rs16840252, and "C" allele of rs5742909 were significantly higher in patients than in controls. Moreover, these alleles also showed significantly higher risk of TAARs (OR = 2.357, 95%CI: 1.584-3.508, p = 0.02; OR = 2.357, 95%CI: 1.584-3.508, p = 0.02; OR = 2.462, 95%CI: 1.619-3.742, p = 0.008; OR = 2.357, 95%CI: 1.584-3.508, p = 0.02; OR = 2.357, 95%CI: 1.584-3.508, p = 0.02, respectively). The present study demonstrated the correlation of CTLA4 gene polymorphism and transfusion reaction, and alleles of 4 CTLA4 SNPs with an increased risk of TAARs were found. It is important to explore the potential immune regulatory mechanism affected by SNPs of costimulatory molecules, and it could predict transfusion reaction occurrence and guide preventive actions.

Investigation of the Correlation between Graves' Ophthalmopathy and CTLA4 Gene Polymorphism.

Graves' disease (GD) is an autoimmune inflammatory disease, and Graves' ophthalmopathy (GO) occurs in 25-50% of patients with GD. Several susceptible genes were identified to be associated with GO in some genetic analysis studies, including the immune regulatory gene CTLA4. We aimed to find out the correlation of CTLA4 gene polymorphism and GO. A total of 42 participants were enrolled in this study, consisting of 22 patients with GO and 20 healthy controls. Chi-square or Fisher's exact test were used to appraise the association between Graves' ophthalmopathy and CTLA4 single nucleotide polymorphisms (SNPs). All regions of CTLA4 including promoter, exon and 3'UTR were investigated. There was no nucleotide substitution in exon 2 and exon 3 of CTLA4 region, and the allele frequencies of CTLA4 polymorphisms had no significant difference between patients with GO and controls. However, the genotype frequency of "TT" genotype in rs733618 significantly differed between patients with GO and healthy controls (OR = 0.421, 95%CI: 0.290-0.611, p = 0.043), and the "CC" and "CT" genotype in rs16840252 were nearly significantly differed in genotype frequency (p = 0.052). Haplotype analysis showed that CTLA4 Crs733618Crs16840252 might increase the risk of GO (OR = 2.375, 95%CI: 1.636-3.448, p = 0.043). In conclusion, CTLA4 Crs733618Crs16840252 was found to be a potential marker for GO, and these haplotypes would be ethnicity-specific. Clinical application of CTLA4 Crs733618Crs16840252 in predicting GO in GD patients may be beneficial.

Human platelet antigens in disease.

Platelets have various functions and participate in primary hemostasis, inflammation, and immune responses. Human platelet antigens (HPAs) are alloantigens expressed on the platelet membrane. Each HPA represent one of six platelet glycoproteins GPIIb, GPIIIa, GPIa, GPIbα, GPIbß, and CD109, and six biallelic systems are grouped. A single nucleotide polymorphism (SNP) in the gene sequence causes a single amino acid substitution of relevant platelet glycoprotein with the exception of HPA-14bw. High-throughput next-generation sequencing-based method has been developed, which enable accurately identification of HPA polymorphisms. The roles of HPA in disease were reviewed. HPAs mediate platelet-microorganism and platelet-malignant cell interactions, and they also participate in pathogenesis of hemorrhagic fever with renal syndrome and infective endocarditis. The exploration of HPA polymorphisms in association with disease susceptibility of individuals will benefit prevention or management of disease.

Sustained or higher levels of growth factors in platelet-rich plasma during 7-day storage.

BACKGROUND: The effectiveness of platelet-rich plasma (PRP) for treating soft tissue injuries is still controversial. Most of PRPs were prepared simply by concentrating in volume and were injected right after preparation in physician offices. Neither platelet count nor growth factors were quantitated in advance. We prepared and stored leukocyte and platelet-rich plasma (L-PRP) by regular separation protocols for blood components in the blood bank. And we investigated the dynamic change of growth factors in the L-PRPs over the period of storage. METHODS: The L-PRPs were prepared by 2-step centrifugation and stored agitatedly at 22 °C for 7 days in the platelet incubator of blood bank. Levels of vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF)-basic, hepatocyte growth factor (HGF), insulin-like growth factor (IGF)-1, platelet derived growth factor (PDGF)-AB, endothelial growth factor (EGF), and transforming growth factor (TGF) over the period of storage were evaluated daily after freeze-thawing to release growth factors from platelet. RESULTS: Compared to original whole blood, platelet concentration, VEGF, FGF-basic, PDGF-AB, EGF, and TGF-beta1 levels of L-PRPs significantly increased after PRP preparation. Both HGF and IGF-1 in L-PRPs remained the original plasma level. Platelet, FGF, and TGF-beta1 concentrations sustained during storage, and concentrations of VEGF, HGF, IGF-1, PDGF-AB, and EGF in L-PRPs increased over the period of storage. CONCLUSIONS: During the storage in blood bank, platelet counts and 7 growth factors sustained or reached higher level than L-PRP obtained on first day. Multiple injections of stored PRPs could become applicable by our protocol.

Human platelet antigens are associated with febrile non-hemolytic transfusion reactions.

BACKGROUND: Febrile non-hemolytic transfusion reaction (FNHTR) is the most common type of transfusion reactions, and it could be reduced by transfusing patients with leukocyte-poor blood products. However, FNHTR still occur in certain patients transfused with leukocyte-poor red blood cell (LPR) products. It is examined whether human platelet antigen (HPA) could be a potential membrane antigen that plays a role in FNHTR. METHODS: A total of 120 inpatient subjects who transfused with LPR (60 in FNHTR group, 60 in control group) were typed for HPA-2, HPA-3, and HPA-15 using sequence specific primer-polymerase chain reaction (SSP-PCR) and electrophoresis. RESULTS: HPA-2 unmatched rate between donors and patients in FNHTR group was 18%, and only 3% unmatched rate was observed in control group (p=0.0082). FNHTR group was further classified according to the imputability. There was a significant difference (p=0.0041) between FNHTR (probable imputability, infection) group and control group, and more significant difference (p=0.0008) was seen between FNHTR (probable imputability, febrile neutropenia) group and control group. CONCLUSIONS: Those results indicated that HPA-2 might play roles on inducing FNHTR in patients suffering from infectious diseases and febrile neutropenia. HPA-2 genotyping between donors and recipients might be worth integrating in pre-transfusion testing to increase transfusion safety.