Objective To investigate the relationship between the levels of peripheral blood iri-sin,myostatin(MSTN),and 25-hydroxyvitamin D[25(OH)D]and the risk and predictive value of sarcopenia in elderly patients with type 2 diabetes mellitus(T2DM)complicated by heart failure.Methods Elderly patients with T2DM complicated by heart failure who visited Xingtai Central Hospi-tal from March 2023 to March 2024 were selected as the study subjects.The sample size was calculated based on the effect size.Patients were divided into modeling set(n=140)and validation set(n=60)at a ratio of 7 to 3 using a simple randomization method.Additionally,according to the occur-rence of sarcopenia,patients in the modeling set were divided into sarcopenia group and non-sar-copenia group.Clinical data and the levels of peripheral blood irisin,MSTN,and 25(OH)D were compared between the modeling and validation sets.The receiver operating characteristic(ROC)curve was used to analyze the predictive efficacy of peripheral blood irisin,MSTN,and 25(OH)D for sarcopenia.Multivariate logistic regression analysis was employed to identify the influencing fac-tors of sarcopenia in elderly patients with T2DM complicated by heart failure,and a relevant predic-tive model was constructed using R software.Results Among 200 elderly patients with T2DM com-plicated by heart failure,71 cases developed sarcopenia,with an incidence rate of 35.50％.The proportion of regular exercise,bone mineral content,and the levels of irisin and 25(OH)D in the sarcopenia group were lower than those in the non-sarcopenia group,while the MSTN level was high-er in the sarcopenia group,and the differences were statistically significant(P＜0.05).The areas under the curve(AUCs)for predicting sarcopenia by irisin,MSTN,and 25(OH)D were 0.878(95％CI,0.812 to 0.927),0.848(95％CI,0.778 to 0.903),and 0.826(95％CI,0.753 to 0.885),respectively.The sensitivities were 74.16％,79.78％,and 88.76％,and the specifici-ties were 74.16％,79.79％,and 88.76％,respectively.The results of multivariate logistic regres-sion analysis showed that lack of exercise habits(OR=2.489,95％CI,1.665 to 3.735),de-creased bone mineral content(OR=2.340,95％CI,1.596 to 3.595),irisin ≤105.44 ng/mL(OR=3.111,95％CI,2.004 to5.147),MSTN＞19.06 μg/L(OR=2.667,95％CI,2.015 to 4.693),and25(OH)D ≤12.23 ng/mL(OR=2.547,95％CI,1.285 to 4.492)were all inde-pendent risk factors for sarcopenia in these patients(P＜0.05).The results of ROC curve analysis showed that the AUCs of the nomogram model for predicting postoperative recurrence in the modeling and validation sets were 0.875 and 0.853,respectively.The Hosmer-Lemeshow test for the model-ing and validation sets showed the following results:x2=0.715,P=0.510;x2=0.651,P=0.568.The calibration curves were basically consistent with the standard curves.The threshold probability range of decision curve analysis(DCA)was 0.1 to 0.9.Within this range,both the modeling and validation sets showed good clinical net benefits.Conclusion Peripheral blood iri-sin,MSTN,and 25(OH)D all have certain predictive values for the occurrence of sarcopenia in elderly patients with T2DM complicated by heart failure.The nomogram model constructed based on irisin,MSTN,and 25(OH)D can provide a quantitative basis for the early screening of sarcopenia in these patients.

OBJECTIVETo construct a retroviral-mediated vector of FLT3 ligand (FL) and express it in human bone marrow stromal cells.

METHODSFL cDNA was inserted into the retroviral vector pLXIN by gene recombination technology. The recombinant plasmid pLFIN was transferred into retrovirus packaging cell line PA317 by lipofectamine, and the positive clones were selected by G418. The mRNA expression in human stromal cells and integration of genome DNA were assayed by reverse transcriptase-polymerase chain reaction (RT-PCR) and genomic DNA-PCR. The expression of FL protein and its biological activities in the culture were investigated by ELISA and mouse bone marrow CFU-GM assay.

RESULTSThe recombinant plasmid pLFIN was successfully constructed. In genome of these transfected target cells, Neo gene and FL gene were integrated, FL mRNA was transcripted and FL protein was expressed at 4.35 ng/ml/24 h. The specific activities of FL in the culture indicated that human bone marrow stromal cells transfected with FL could significantly express FL in vitro.

CONCLUSIONThe retroviral-mediated FL gene was expressed in bone marrow stromal cells and the biological activities of FL were detectable in the supernatant of the transfected cells. These results provide a basis for studies on hematopoietic regulation by gene transfected bone marrow stromal cells.

3T3 Cells ; Animals ; Bone Marrow Cells ; cytology ; metabolism ; Cell Line ; Colony-Forming Units Assay ; Enzyme-Linked Immunosorbent Assay ; Gene Expression ; Genetic Vectors ; genetics ; Humans ; Membrane Proteins ; genetics ; metabolism ; Mice ; RNA, Messenger ; genetics ; metabolism ; Retroviridae ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Stromal Cells ; cytology ; metabolism ; Transfection

The generation of large quantities of novel human T cell clones ex vivo would make a wide range of gene-and immuno-therapies for tumor and AIDS possibly. Although it is well established that T cells are derived from CD34(+) cells, the involvement of thymic fragments from either human or murine fetus makes the in vitro T cell perliferation very cumbersome. In this report, cord blood mononuclear cells were used as accessory cells to support the differentiation of CD34(+) cells into naive T cells stimulated with SCF and IL-2. CD4(+) and CD8(+) T cells, under the cultural conditions, were continuously produced in vitro at least over a period of 3 weeks and their ratios changed gradually. CD4/CD8 double positive T cells and RAG-2 gene were existed, and RAG-2 gene, reponsible for TCR rearrangement, was expressed during the cell proliferation. Our study presents a simple culture system in vitro to acquire large quantities of naive T cell clones.

Transfer of drug resistance genes into hematopoietic cells is an attractive approach to protect hematopoietic system from the toxic effects by chemotherapeutic agents in cancer patients. In this study, transduction of mdr-1 in combination with dihydrofolate reductase (dhfr) gene was performed, and the expression of exogenous genes and chemoprotection capacity in mouse bone marrow cells were observed. The results showed that approximately 15% of bone marrow cells transfected with the retroviral vector expressed mdr-1 as assayed by flow cytometry. Gene transfer resulted in about 0.9 - 13 fold and 0.5 - 2.6 fold increase in the resistance of CFU-GM to taxol and methotrexate in vitro, respectively (P < 0.05). Moreover, seven months after transplantation to syngeneic mice with mdr-1 and dhfr-transfected bone marrow cells, peripheral blood cells in recipients were still positive for gp170 as evaluated by FACS as well as for mdr-1 and dhfr by PCR amplification. These results indicate that hematopoietic progenitors can be transfected by retrovirus containing mdr-1 and dhfr genes, and that functional drug resistance accompanies their expressions. Furthermore, genetic chimerism might exist in hematopoietic stem cells. In conclusion, transfer and expression of mdr-1 and dhfr genes in bone marrow cells might be applicable in gene therapy research in cancer patients.