Abstract We have previously demonstrated the cardioprotective effects of exosomes derived from mesenchymal stem cells (MSCs). It is well known that the activation of Akt is involved in stem cell-induced cardioprotection. In the present study, we investigated whether exosomes released from Akt-overexpressing MSCs showed a beneficial effect on cardioprotection and angiogenesis. MSCs were collected from human umbilical cord (hucMSCs), and Akt was transfected into hucMSCs (Akt-hucMSCs) by using an adenovirus transfection system. Exosomes were isolated from control hucMSCs (Exo) and Akt-hucMSCs (Akt-Exo). An acute myocardial infarction model was created by ligation of the left anterior decedent coronary artery (LAD) in rats. Various source exosomes (400 µg of protein) were infused via the tail vein immediately after LAD ligation. The cardiac function was evaluated by using echocardiography after different treatments for 1 and 5 weeks, respectively. Endothelial cell proliferation, migration, and tube-like structure formation, as well as chick allantoic membrane assay, were used to evaluate the angiogenetic effects of Akt-Exo. The results indicated that cardiac function was significantly improved in the animals treated with Akt-Exo. In addition, Akt-Exo significantly accelerated endothelial cell proliferation and migration, tube-like structure formation in vitro, and blood vessel formation in vivo. The expression of platelet-derived growth factor D (PDGF-D) was significantly upregulated in Akt-Exo. However, the angiogenesis was abrogated in endothelial cells treated with the exosomes obtained from MSCs transfected with PDGF-D-siRNA. Our studies suggest that exosomes obtained from Akt-modified hucMSCs are more effective in myocardial infarction therapy through promoting angiogenesis. PDGF-D plays an important role in Akt-Exo-mediated angiogenesis.
Cellular immunotherapy exploits the capacity of the human immune system in self-protection and surveillance to achieve the anti-tumor effects. Natural killer (NK) cells are lymphocytes of innate immune system and they display a unique inherent ability to identify and eliminate tumor cells. In this review, we first introduce the basic characteristics of NK cells in the physiological and pathological milieus, followed by a discussion of their effector function and immunosuppression in the tumor microenvironment. Clinical strategies and reports regarding NK cellular therapy are analyzed in the context of tumor treatment, especially against solid tumors. Given the widely studied T-cell therapy in the recent years, particularly the chimeric antigen receptor (CAR) T-cell therapy, we compare the technical features of NK- and T-cell based tumor therapies at the clinical front. Finally, the technical challenges and potential solutions for both T and NK cell-based immunotherapies in treating tumor malignancies are delineated. By overviewing its clinical applications, we envision the NK-cell based immunotherapy as an up-and-comer in cancer therapeutics.
Objective: To investigate whether the characteristics of human umbilical cord mesenchymal stem cells(hucMSCs) labeled with CM-Dil will be changed;and if CM-Dil labled hucMSCs can be traced in vivo.Methods: HucMSCs were isolated from human umbilical cord.The hucMSCs of passage 3 were selected for labeling in vitro.Using cell counting method to analyse growth curves of CM-Dil labeled hucMSCs;observed CM-Dil labeled hucMSCs in vivo with imaging instrument.Results: The efficiency of CM-Dil dye-labeled hucMSCs was high.The proliferation of labeled hucMSCs had no obvious change.CM-Dil dye-labeled hucMSCs can be observed in vivo.Conclusion: CM-Dil labeled hucMSCs can be traced in vivo.
Bats are important reservoirs for many kinds of emerging zoonotic viruses. In order to explore potential pathogens carried by bats and trace the source of adenovirus outbreaks on the southeastern coast of China, we took pharyngeal and anal swabs from a total of 552 bats (Rhinolophus pusillus) collected from various areas of Chinese southeastern coast. Adenoviruses were identified in 36 out of the 552 samples (6.5%) . Complete genome sequences of two adenovirus isolations from Vero E6 cells were obtained, which were further validated as identical strains via next-generation sequencing and were named Bat-Advcxc6. The cell culture inoculated with the two samples exhibited remarkable cytopathic changes. The full genome has 37,315 bp and owns 29 open reading frames. Phylogenetic analyses confirmed that Bat-Advcxc6 represented a novel bat adenovirus species in the genus Mastadenovirus. Transmission electron microgram showed clear virus particles. Bat-Advcxc6 shared similar characteristics of G + C contents with Bat mastadenovirus WIV11 (Bat mastadenovirus C) found in China in 2016, but differed from this serotype due to a <75% similarity with DNA polymerase amino acid sequences in WIV11. As it is a newly found adenovirus strain according to the international classification criteria, further analyses of virus dynamics, epithelial invasion, and immunization assays are required to explore its potential threats of cross-species transmission.
Objective: To construct green fluorescence protein(GFP)labeled recombinant adenovirus vector carrying TNFα-Tumstatin fusion gene and transfect it into hUCMSCs.Methods: The amplification products of TNFα-Tumstatin by PCR with a pair of primers with BglⅡand HindⅢ restriction endonuclease sites and signal peptide sequence of GM-CSF were subcloned into shuttle plasmid pAdtrack-CMV after digesting with BglⅡand HindⅢ.The resultant plasmid,after linearized by digesting with restriction endonuclease PmeⅠ,was transformed into E.coli.BJ5183 that had been transformed by adenoviral backbone plasmid pAdEasy-1.Recombinant plasmid were obtained by alternation of kanamycin and then confirmed by PCR and restriction endonuclease analysis.The adenovirus was packaged and propagated in human embryonal kidney cells(293A cells)after being linearized by digesting with restriction endonuclease PacⅠand then was transfected into hUCMSCs.Fusion gene expression in infected hUCMSCs was detected by RT-PCR. Results: The results of PCR and restriction endonuclease assay indicated that target gene was inserted into recombinant adenovirus vector successfully.The sequence of fusion gene was the same as that of designed fragments.hUCMSCs infected by the recombinant adenovirus expressed GFP and fusion gene which could be demonstrated by fluorescence microscope and RT-PCR respectively. Conclusion: Recombinant adenovirus vector containing TNFα-Tumstatin has been constructed successfully and could transfected hUCMSCs efficiently,which laid a foundation for further investigation of anti-tumor effect of hUCMSCs modified with TNFα-Tumstatin.
Metastasis is the major cause of death in breast cancer patients. Although the strategies targeting metastasis have promoted survival, the underlying mechanisms still remain unclear. In this study, we used microarray data of primary breast tumor, tumor derived from bone and liver, and skin metastatic tissue, to identify the key genes and pathways that are involved in metastasis in breast cancer.We first calculated the differentially expressed genes (DEGs) between three metastatic tissues and primary tumor tissue, and then used it to perform Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Further, we analyzed the correlation of genes enriched in GO terms and KEGG pathways with survival of breast cancer patients. To identify the key genes and pathways associated with metastasis, we overlapped the DEGs and KEGG pathways. In our in vitro experiments, we knocked down the key gene, ERLIN2, and detected the PI3K expression in tumor cells to evaluate their effect on tumor metastasis.We identified six genes (ALOX15, COL4A6, LMB13, MTAP, PLA2G4A, TAT) that correlated with survival. Seven key genes (SNRPN, ARNT2, HDGFRP3, ERO1LB, ERLIN2, YBX2, EBF4) and seven signaling pathways (metabolic pathways, phagosome pathway, PI3K-AKT signaling pathway, focal adhesion, ECM-receptor interaction, pancreatic secretion, human papillomavirus infection) associated with metastasis were also identified. Our in vitro experiments revealed that ERLIN2 was highly expressed in MDA-MB231 cells compared to MCF-7 cells. Moreover, knockdown of ERLIN2 increased apoptosis, while inhibiting the proliferation, invasion, and migration ability of breast cancer cells. The PI3K/AKT signaling pathway was also found to be highly expressed in MDA-MB231 cells.Our results reveal the key genes and signaling pathways that contribute to metastasis, and highlight that strategic targeting of ENLIN2 and PI3K/AKT signaling pathways could inhibit metastasis of breast cancer.
Background. The prognostic role of CRP (C-reactive protein) in gynecological tumors has been previously reported in individual studies, but whether CRP can be used as a separate potential prognostic factor has not been systematically reviewed. The purpose of this research is to determine if there is a link between CRP levels and the prognosis of gynecological cancer patients. Methods. A systematic search was carried out to find the literature evaluating the predictive role of CRP in the prognosis of gynecological cancer patients. For the purpose of determining the relationship between CRP and clinicopathological characteristics, the pooled odds ratio (OR) was calculated. A hazard ratio (HR) with a 95% confidence interval (CI) was used to determine differences in overall survival (OS), disease-free survival (DFS), or progression-free survival (PFS) between patients with low and high CRP levels. Results. A total of 19 studies, including 4062 patients, were analyzed retrospectively. The FIGO stage was related to the CRP level (OR = 0.43, 95% CI: 0.19–1.00). Age, lymph node metastasis, and histological grade were not associated with CRP level (OR = 0.93, 95% CI: 0.69–1.25; OR = 0.91, 95% CI: 0.65–1.28; OR = 0.74, 95% CI: 0.52–1.05). Worse OS (HR = 1.40, 95% CI: 1.23–1.57), DFS (HR = 1.20, 95% CI: 1.12–1.28), and PFS (HR = 1.57, 95%CI: 1.23–1.91) were associated with elevated CRP levels, as shown by the pooled results. Subgroup analysis was performed according to cancer type (endometrial cancer: HR = 1.15, 95% CI: 1.02–1.28; ovarian cancer: HR = 1.67, 95% CI: 1.03–2.31; cervical cancer: HR = 1.42, 95% CI: 1.19–1.64), multivariate value (HR = 1.22, 95% CI: 1.10–1.33), and age (HR = 1.50, 95% CI: 1.28–1.72). Significant correlations were observed between CRP and OS. Conclusions. CRP may be utilized as a prognostic indicator for a variety of gynecologic malignancies, including cervical cancer, ovarian cancer, endometrial cancer, and vulvar cancer.
Abstract Precocious puberty is a common phenomenon in crab breeding that seriously reduces the economic benefits for crab farmers. To address this problem, this study aimed to explore the potential functions of both methylation and hydroxymethylation of testis rRNA genes with respect to precocious puberty in Eriocheir sinensis . The results showed that the rRNA genes in normally developing testes of E. sinensis had low levels of methylation and high levels of hydroxymethylation; however, although methylation levels were similar, the level of hydroxymethylation in precocious testes was lower than normal. Highly significant differences ( P < 0.01 ) in the hydroxymethylation of the 18S and 28S rRNA genes were found between precocious and normal testes. Our results suggested that both the 18S and 28S rRNA genes, which are normally downregulated by hypo-hydroxymethylation, might be involved in the process of precocious puberty. Our results also implied that hydroxymethylation of the 18S and 28S rRNA genes might be used as an important epigenetic molecular marker to evaluate economically significant potential for growth and breeding in this species.
To investigate multitissue engraftment of human primitive hematopoietic cells and their differentiation in goats, human CD34 + Lin − cord blood cells transduced with a GFP vector were transplanted into fetal goats at 45–55 days of gestation. GFP + cells were detected in hematopoietic and nonhematopoietic organs including blood, bone marrow, spleen, liver, kidney, muscle, lung, and heart of the recipient goats (1.2–36% of all cells examined). We identified human β2 microglobulin-positive cells in multiple tissues. GFP + cells sorted from the perfused liver of a transplant goat showed human insulin-like growth factor 1 gene sequences, indicating that the engrafted GFP + cells were of human origin. A substantial fraction of cells engrafted in goat livers expressed the human hepatocyte-specific antigen, proliferating cell nuclear antigen, albumin, hepatocyte nuclear factor, and GFP. DNA content analysis showed no evidence for cellular fusion. Long-term engraftment of GFP + cells could be detected in the blood of goats for up to 2 yr. Microarray analysis indicated that human genes from a variety of functional categories were expressed in chimeric livers and blood. The human/goat xenotransplant model provides a unique system to study the kinetics of hematopoietic stem cell engraftment, gene expression, and possible stem cell plasticity under noninjured conditions.
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