Molecular polymorphism Analysis on CD36 Deficiency among Platelet Blood Donors in Shenzhen / 中国实验血液学杂志

OBJECTIVE@#To analyze the molecular polymorphisms of CD36 among 58 blood donors with CD36 deficiency and compare with CD36 positive controls.@*METHODS@#A total of 58 donors with CD36 deficiency during a screening conducted in the laboratory from September 2019 to December 2020 were enrolled as the test group, including 39 males and 19 females, while 120 platelet donors with CD36 positive were randomly selected as the controls, including 76 males and 44 females. All of the subjects were Han nationality. The PCR-SBT method was used to detect coding region of CD36 gene, and molecular mutations were compared with those CD36 positive controls.@*RESULTS@#Among the 58 donors with CD36 deficiency, mutations appears in 32 individuals. The detection rate for type I was 71.43% (5/7), and type II was 51.92% (27/52), while among the 120 controls, mutations appears in 12 donors (10%). In the CD36 antigen-deficient donors, 16 variations were found, in which 329-330 del AC with the highest frequency accounted for 20.69%, followed by 1228-1239 del ATTGTGCCTATT(15.52%) and 1156 C>T(10.34%). Two variations, 198-205 del GATCTTTG and 220 C>T, led to premature termination of translation; four mutations, 329-330 del AC, 560 ins T, 1011-1049 39bp dupl and 1343-1344 ins TCTT, caused translation frame shift; 1228-1239 del ATTGTGCCTATT led to deletion of four amino acids (Ile-Val-Pro-Ile) at sites 410-413 of the peptide chain. The 1140 T>A and 1275 G>A were synonymous mutations, and the other 7 mutations resulted in the substitution of single nucleotide. The platelet expression in the donors of CD36 positive with 329-330 del AC or 1228-1239 del ATTGTGCCTATT mutation (heterozygote) was lower than those CD36 positive individuals without mutations (homozygote).@*CONCLUSION@#Multiple gene mutations in the CD36 coding region may cause CD36 deficiency, and the heterozygous individuals with mutations may lead to CD36 antigen reduction or deletion. Mutation is not detected in 44.83% of CD36 deficient individuals, there may be some other reasons for the CD36 antigen deficiency.

KLF9 regulates hepatic lipid metabolism via inducing CD36 expression / 生理学报

The development of nonalcoholic fatty liver disease (NAFLD) is closely related to the fatty acid (FA) uptake. This study aimed to investigate the effect of Krüppel-like factor 9 (KLF9) on CD36 (typical fatty acid translocase), hepatocellular lipid metabolism as well as the development and progression of nonalcoholic fatty liver. High-fat diet-induced obese C57BL/6J mice and db/db mice were used to test the expression levels of Klf9 and Cd36 in the livers. The primary hepatocytes were isolated from C57BL/6J mice, treated with Ad-GFP, Ad-Klf9, Ad-shCtrl or Ad-shKlf9, and then incubated with oleic acid and palmitic acid for 24 h. Liver-specific knockout of Klf9 mice were established. The protein levels and relative mRNA levels were examined by Western blot and real-time PCR, respectively. Triglyceride content was determined by using an assay kit. Lipid content was determined by Oil Red O staining. The results showed that: (1) Klf9 expression levels were increased in the livers of high-fat diet-induced obese mice and db/db mice, compared to their respective control mice. (2) Adenovirus-mediated overexpression of Klf9 in primary hepatocytes increased Cd36 expression and cellular triglyceride contents. (3) In contrast, adenovirus-mediated knockdown of Klf9 expression in primary hepatocytes by Ad-shKlf9 decreased Cd36 expression and cellular triglyceride contents. (4) Finally, Klf9 deficiency decreased liver Cd36 expression and alleviated fatty liver phenotype of high-fat diet-induced obese mice. These results suggest that KLF9 can regulate hepatic lipid metabolism and development of NAFLD by promoting the expression of CD36.

Platelet Rich Plasma (PRP) Produces an Atherofibrotic Histophenotype During Craniofacial Bone Repair Due to Changes of Immunohistochemical Expression of Erk1/2, p38α /β, Adiponectin and Elevated Presence of Cells Exhibiting B-scavenger Receptor (CD36+)

Abstract The platelet-extracellular matrix interaction in platelet rich plasma (PRP) through thrombospondin receptor-CD36 induces the secretion of growth factors responsible for cellular proliferation and differentiation during the repair process. Since CD36 also acts as a class B-scavenger-receptor for development of foam-like cells and mitogen-activated kinases, such as Erk1/2 and p38α/β, are important proteins activated by platelet growth factor, the aim of this study was to evaluate the immunohistochemical presence of CD36, Erk1/2, p38α/β during the bone repair treated and non-treated with PRP and to compare these results with the histomorphometry of repair. Simultaneously, the immunopresence of adiponectin was analyzed, which may contribute to osteogenesis at the same time it inhibits fibrosis and impairs adipogenesis and foam cell formation in the medullary area. An artificial bone defect measuring 5×1 mm was produced in the calvaria of 56 Wistar rats. The defects were randomly treated with autograft, autograft+PRP, PRP alone and sham. The animals were euthanized at 2 and 6 weeks post-surgery. Data were analyzed by ANOVA followed by non-parametric test Student Newman-Keuls (p<0.05) for histomorphometric and immunohistochemical interpretation. The results revealed that in specimens that received PRP the immunopositivity for Erk1/2, p38α/β and CD36 proteins increased significantly while the immunohistochemical expression of adiponectin decreased simultaneously. There was also an accentuated reduction of bone matrix deposition and increase of the medullary area represented by fibrosis and/or presence of foam-like cells, which exhibited immunophenotype CD36+adiponectin. The findings of this study suggest that PRP acted as an inhibitor of osteogenesis during the craniofacial bone repair and induced a pathological condition that mimics an atherofibrotic condition.

Resumo A interação da matriz extracelular-plaquetas no plasma rico em plaquetas (PRP) através de receptor trombospondina CD36 induz a secreção de fatores de crescimento responsáveis pela proliferação e diferenciação celular durante o processo de reparo. Uma vez que o CD36 também age como receptor scavenger de classe B para o desenvolvimento de células do tipo espuma, e as quinases ativadas por mitógenos, tais como ERK1/2 e p38α/β, são importantes proteínas ativadas por fator de crescimento das plaquetas, o objetivo deste estudo foi avaliar a presença imunoistoquímica de CD36, ERK1/2, p38α/β durante o reparo ósseo tratado e não-tratado com PRP e comparar estes resultados com a histomorfometria do reparo. Simultaneamente, analisou-se a imunopresença da adiponectina, que pode contribuir para osteogênese ao mesmo tempo que inibe a fibrose e prejudica a formação de células tipo espuma/xantomatosas na área medular. Um defeito artificial de osso medindo 5×1 mm foi produzido na calvária de 56 ratos Wistar. Os defeitos foram tratados aleatoriamente com auto-enxerto, enxerto autógeno+PRP, PRP apenas e sham. Os animais foram sacrificados 2 e 6 semanas pós-cirurgia. Os dados foram examinados por meio de ANOVA, seguido pelo teste não-paramétrico Student Newman-Keuls (p<0,05) para a interpretação histomorfométrica e imunoistoquímica. Os resultados revelaram que as amostras que receberam PRP aumentaram significativamente a imunopositividade para as proteínas ERK1/2, p38α/β e CD36, simultaneamente à diminuição de expressão imunoistoquímica da adiponectina. Houve também expressiva redução de deposição de matriz óssea e aumento da área medular representada por fibrose e/ou presença de células do tipo espuma que apresentaram imunofenótipo CD36 + adiponectina. Estes resultados sugerem que o PRP atuou como um inibidor da osteogênese durante o reparo ósseo craniofacial e induziu uma condição patológica que mimetiza uma condição aterofibrótica.

Topology of Scavenger Receptor Class B Type I (SR-BI) on Brush Border Membrane

Both hydropathy plot and in vitro translation results predict the topology of SR-BI; the receptor is an integral membrane protein of 509 amino acids, consisting of a short cytoplasmic N-terminus of 9 amino acids followed by a first transmembrane domain of 22 amino acids, the extracellular domain of 408 amino acids, the second transmembrane domain of 22 amino acids, and the cytoplasmic C-terminus of 47 amino acids. The immunoblot of rBBMV in the presence or absence of pAb589 peptide antigen (the C-terminal 22 amino acid residues of SR-BI) confirmed that the bands at apparent molecular weight of 140 and 210 kDa are SR-BI related protein which might be multimeric forms of SR-BI. 125I apo A-I overlay analysis showed that SR-BI can bind to its ligand, apo A-I, only when it is thoroughly matured - glycosylated and dimerized. The antibody which was generated against extracellular domain of SR-BI (pAb230) not only prevented 125I-labeled apo A-I from binding to 140 kDa band but also inhibited the esterified cholesterol uptake of rabbit BBMV with its IC50 value of 40 microgram/ml of IgG. In contrast, the antibody generated against the C-terminal domain of SR-BI (pAb589) did not show any effect either on cholesterol uptake of rabbit BBMV or 125I-labeled apo A-I binding to 140 kDa band. Overall results show that the ligand binding site of SR-BI in rabbit BBMV is located in extracellular domain, and SR-BI is only functional when it is part of dimeric forms which rationalize the previously found cooperative nature of the binding interaction and maybe a fundamental finding towards the so far poorly understood mechanism of SR-BI function.