BACKGROUND:Spleen has the functions of blood storage,hematopoiesis,and immunity.With the increase of age,the structural degeneration and functional decline of spleen lead to the impairment of immune system function,thus accelerating the aging process of the body.The treatment of spleen aging in tree shrews with highly active umbilical cord mesenchymal stem cells has not been reported. OBJECTIVE:To explore the intervention effect and mechanism of highly active umbilical cord mesenchymal stem cells on spleen aging in tree shrews. METHODS:Highly active umbilical cord mesenchymal stem cells were isolated,cultured,and obtained from the umbilical cord tissue of newborn tree shrews by caesarean section.The differentiation abilities of adipogenesis,osteogenesis,and chondrogenesis were detected by three-line differentiation kit.Cell cycle and surface markers were detected by flow cytometry.The second generation of highly active umbilical cord mesenchymal stem cells were transfected with Genechem Green Fluorescent Protein with infection complex values of 100,120,140,160,180,and 200,respectively,to screen the best transfection conditions.After transfection,the fourth generation of highly active umbilical cord mesenchymal stem cells was injected into the tail vein of tree shrews in the elderly treatment group.The young control group and the aged model group were not given special treatment.After 4 months of treatment,the spleen tissue was taken and the structure of the spleen was observed by hematoxylin-eosin staining.β-Galactosidase staining was used to detect the activity of aging-related galactosidase.Immunohistochemical staining was used to detect the expression levels of p21 and p53 proteins.Ki67 and PCNA immunofluorescence staining was used to detect cell proliferation activity.Immunofluorescence staining was used to detect the expression levels of spleen autophagy protein molecules Beclin 1 and APG5L/ATG5.Reactive oxygen species fluorescence staining was used to detect the content of reactive oxygen species in spleen tissue.CD3 immunofluorescence staining was used to detect the change of the proportion of total T lymphocytes.The secretion levels of interleukin 1β and transforming growth factor β1 in spleen were detected by enzyme linked immunosorbent assay.The distribution of highly active umbilical cord mesenchymal stem cells labeled with green fluorescent protein in spleen tissue was observed by DAPI double staining of nucleus. RESULTS AND CONCLUSION:(1)Highly active umbilical cord mesenchymal stem cells grew in a short spindle shape with fish-like growth,with a large proportion of G0/G1 phase,and had the potential to differentiate into adipogenesis,osteogenesis,and chondrogenesis.(2)Multiplicity of infection=140 and transfection for 72 hours were the best conditions for labeling tree shrews highly active umbilical cord mesenchymal stem cells with Genechem Green Fluorescent Protein.(3)Compared with the aged model group,in the aged treatment group,the spleen tissue cells of tree shrews were arranged closely,and the area of white pulp was increased(P<0.01);the boundary between red pulp and white pulp was clear;the proportion of germinal centers did not show statistically significant difference(P>0.05).The activity level of galactosidase related to spleen tissue aging was decreased(P<0.001),and the expression levels of aging protein molecules p21 and p53 were down-regulated(P<0.001).The expression levels of proliferation-related molecules Ki67 and PCNA were up-regulated(P<0.001,P<0.05);expression levels of autophagy-related molecules Beclin 1 and APG5L/ATG5 were up-regulated(P<0.001),and the content of reactive oxygen species decreased(P<0.001),and the proportion of CD3+T cells increased(P<0.05).The secretion level of interleukin 1β in the aging-related secretion phenotype decreased(P<0.001);no significant difference was found in transforming growth factor β1 level(P>0.05).Compared with the young control group,the above indexes were significantly different in the elderly treatment group(P<0.05).(4)Green fluorescent cells labeled with green fluorescent protein were observed in spleen tissue of tree shrews the elderly treatment group by frozen tissue section observation.The results show that intravenous infusion of highly active umbilical cord mesenchymal stem cells can migrate to spleen tissue,inhibit the production of reactive oxygen species,down-regulate the expression of aging-related proteins,induce autophagy,promote cell proliferation,reduce chronic inflammation,and then improve the structure and function of spleen tissue.

BACKGROUND:Despite a series of clinical treatment measures,the treatment of pulmonary fibrosis still faces challenges.In recent years,mesenchymal stem cells and their extracellular vesicles have attracted extensive attention as an emerging therapeutic strategy and are considered to be a promising means of treating pulmonary fibrosis. OBJECTIVE:To systematically review the application of mesenchymal stem cells and their extracellular vesicles in the treatment of pulmonary fibrosis,to comprehensively understand their therapeutic mechanism,efficacy evaluation and problems,and provide reference and guidance for further research and clinical application in the future. METHODS:Using Chinese and English search terms"mesenchymal stem cells","mesenchymal stem cell extracellular vesicles","pulmonary fibrosis",we searched the CNKI and PubMed electronic journal databases.By means of manual reading and eliminating duplicate articles,112 articles were selected,but 58 Chinese and English articles were finally included for summary. RESULTS AND CONCLUSION:(1)Mesenchymal stem cells and their extracellular vesicles have shown great potential in the treatment of pulmonary fibrosis,such as regulating inflammatory responses,inhibiting fibroblast proliferation,and promoting damaged tissue repair.Preliminary results from clinical trials have also shown some effects of the treatment,including improved lung function and quality of life in patients.(2)However,mesenchymal stem cells and extracellular vesicles in the treatment of pulmonary fibrosis still face some challenges.During treatment,technical challenges such as cell migration and intrachistological localization need to be addressed for it to accurately reach the damaged lung tissue.Furthermore,its long-term safety also needs to be further studied and improved.For translational medicine development,standardized procedures such as cell collection,cell isolation,cell culture,cell harvesting,and cell identification need to be refined.(3)Despite these challenges,through the joint efforts of scientific researchers and medical personnel,these problems are expected to be gradually solved.In the future,we can further improve treatment outcomes by optimizing treatment regimens and exploring individualized treatments.At the same time,in-depth research on the therapeutic mechanism of stem cells and their extracellular vesicles is expected to develop more efficient and safe therapeutic strategies.

Objective:To elucidate the change of whole genome expression profile for the effect of melatonin on radiation-induced intestinal injury in mice.Methods:C57BL/6J male mice were administrated with melatonin at 10 mg/kg body weight by intraperitoneal injection once a day for five consecutive days before abdominal irradiation with 14 Gy of γ-rays. Small intestines were harvested 3 d after radiation. GO annotation and KEGG pathway of the differential genes involved in small intestine were explored by DNA microarray analysis.Results:Compared with the control group, 584 differential genes were up-regulated and 538 differential genes were down-regulated for administration group pre-irradiation. The overlapping differential genes were selected from the irradiated mice and the administrated mice pre-irradiation. There were 324 up-regulated genes and 246 down-regulated genes unique to the administrated mice pre-irradiation. GO annotation analysis of the differential genes indicated that the top 15 significantly enriched biological processes for the administrated mice pre-irradiation mainly included autophagosome assembly (GO: 0000045), autophagosome organization (GO: 1905037) and regulation of acute inflammatory response (GO: 0002673). The genes ATG12, ATG16L2 and AMBRA1 were involved in autophagosome assembly and autophagosome organization. The genes C3, CPN1, CD55, CFP, CNR1, C1QA, C2 and CREB3L3 were involved in the regulation of acute inflammation response. KEGG pathway analysis of the differential genes involved indicated that the top 15 significantly enriched pathways for the administrated mice pre-irradiation mainly included O-glycan biosynthesis (hsa00512), glycosphingolipid biosynthesis (hsa00603), ECM-receptor interaction (hsa04512) and biosynthesis of unsaturated fatty acids (hsa01040). qRT-PCR verification showed that the expressions of ATG12 and ATG16L2 genes involved in autophagy for the administrated mice pre-irradiation increased significantly compared with the irradiated mice ( t=2.40, 4.35, P<0.05). Conclusions:The differential genes related with the biological process of autophagy, acute inflammatory response and the pathway of unsaturated fatty acid biosynthesis might be involved in the effect of melatonin on radiation-induced intestinal injury.

OBJECTIVE: To analyze variants of TSC1 and TSC2 genes in a Chinese patient with tuberous sclerosis complex (TSC). METHODS: Peripheral blood samples were collected from the patient and her parents with informed consent. Following extraction of genomic DNA, potential variants of the TSC1 and TSC2 genes was detected by using targeted capture next-generation sequencing (NGS) and Sanger sequencing. RESULTS: The patient was found to harbor a de novo mosaicism variant c.3295_3298delG (Val1100CysfsTer3) of the TSC2 gene, with the proportion of the mutant allele determined as 13.4%, which was confirmed by Sanger sequencing. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the c.3295_3298delG (Val1100CysfsTer3) variant was predicted to be pathogenic (PVS1+PS2+PM2). CONCLUSION The mosaicism heterozygous variant of c.3295_3298delG of the TSC2 gene, as detected by both NGS and Sanger sequencing, probably underlay the TSC2 in this patient.
Female ; Humans ; Mosaicism ; Mutation ; Tuberous Sclerosis/genetics* ; Tuberous Sclerosis Complex 1 Protein/genetics* ; Tuberous Sclerosis Complex 2 Protein/genetics*

Objective To analyze the survival and toxicity after concurrent chemoradiotherapy in patients of different ages with stage Ⅳ non-small cell lung cancer (NSCLC).Methods Clinical data of 282 NSCLC patients in two prospective studies were retrospectively analyzed,who completed the protocol (at least 2 cycles of chemotherapy and thoracic radiation doses of ≥36 Gy).Among them,44 patients were assigned into in the young group (≤ 45 years old),161 patients in the middle-age group (46-64 years old) and 77 patients in the elderly group (≥ 65 years old).The clinical characteristics of patients among different groups were analyzed by x2 test.The overall survival (OS) was calculated by Kaplan-Meier method.Stratified analysis was performed by Log-rank test.Multi-factor prognosis analysis was conducted by Cox's proportional hazards regression model.Results The incidence of NSCLC in the male patients in the elderly group was higher than that in the middle-age and young groups.The 1-,2-,3-and 5-year OS did not significantly differ among different groups (P=0.810).The OS did not significantly differ among patients of the same gender,pathological type,T stage,N stage,metastasis status,same chemotherapy cycle,primary tumor dose and comprehensive treatment and short-term response (all P＞0.05).The incidence of adverse events did not considerably differ among different groups.Multivariate analysis demonstrated that age was not an independent factor for survival (P＞ O.05).Conclusion Patients of different ages with stage Ⅳ NSCLC obtain similar survival benefits and adverse events after concurrent chemoradiotherapy.

Objective To investigate the impact of the changes of posttreatment karnofsky performance status (KPSpost) on the overall survival (OS) for patients with stage Ⅳ non-small cell lung cancer (NSCLC) underwent concurrent chemoradiation.Methods A total of 279 patients (male 198 and female 81) with histological confirmed stage Ⅳ NSCLC were enrolled in this study with a median age of 58 years old (range 22 to 80 years old).There were 166 cases of squamous carcinoma,87 cases of adenocarcinoma,and 22 cases of unclassified carcinoma,respectively.All enrolled patients received more than 2 cycles of chemotherapy and more than 36 Gy of concurrent radiotherapy.Kaplan-Meier method and Log-rank test were applied to evaluate OS.Multivariate analyses were carried out by the Cox proportionalhazard model.Chi-square test and logistic regression analysis were used to explore the related factors of KPSpost.Results There were 198 patients with improved KPSpost and 81 patients with decreased KPSpost,respectively.Univariate and multivariate analyses indicated that the improvement of KPSpost was associated with longer OS.Logistic regression analysis showed that the improvement of KPSpost was positively related with treatment of more than 4-6 cycles chemotherapy concurrent with over 63 Gy radiation to primary tumor.The improvement of KPSpost also correlated positively with disease control rate (DCR),but negatively with PLT toxicity and radiation esophagitis.Conclusions KPSpost was an independent prognostic factor of OS for patients with stage Ⅳ NSCLC underwent concurrent chemoradiation.Chemotherapy of 4-6 cycles and concurrent over 63 Gy radiotherapy dose to primary tumor,as well as DCR were positive factors for KPSpost improvement.However,stage 3-4 PLT toxicities and radiation esophagitis decreased the KPSpost.

In recent years, the incidence of hematological malignancies has increased year by year, although with the progress of chemotherapy, targeted therapy and hematopoietic stem cell transplantation, the efficacy of some types of lymphoma and leukemia has been greatly improved.However, the prognosis is still poor and the fatality rate is high.Finding new strategies to improve the curative effect has become an urgent problem to be solved.Oncolytic virus can promote anti-tumor response by selectively destroying tumor cells and inducing specific anti-tumor immune response.At present, oncolytic virus has become an effective strategy for the treatment of a variety of malignant tumors.A lot of breakthrough progress has also been made in the treatment of hematological malignant tumors.In this paper, the latest research progress of oncolytic virus in the treatment of hematological malignant tumors in recent years is reviewed.

Objective: To explore the prognostic relationship between initial absolute lymphocyte count(ALC) of peripheral blood and primary immune thrombocytopenia (ITP) in children, in order to provide basis for judging the prognosis and treatment of ITP in children. Methods: Clinical data of 166 children with primary ITP in children admi-tted at the Affiliated Hospital of Guizhou Medical University from January 2014 to March 2018 were analyzed retrospectively, and they were followed up by clinic and telephone, the prognostic factors (gender, age, ethnicity, inducement, bleeding, initial ALC, platelet count and treatment) were statistically analyzed, and the relationship between ALC of peripheral blood and the prognosis of children with ITP was observed. Results: Of 166 children with ITP, 89 cases (53.6%) had remission within 3 months, 18 cases (10.8%) within 3-12 months, 20 cases (12.0%) within 1-4 years, a total of 39 cases (23.5%) were refractory (no remission in 1-10 years), the remission rate within 1 year was 64.5%, the total remission rate was 76.4%.ALC of remission cases was (4.58±2.87)×10⁹/L within 3 months, (4.47±2.04)×10⁹/L within 3-12 months, and (2.86±1.61)×10⁹/L within 1- 4 years.Thirty-nine cases (23.5%) were refractory(no remission in 1-10 years), ALC of them was (2.07±0.98)×10⁹/L, and there were significant differences among different groups (F= 12.06, P<0.01). Univariate analysis showed that age (F=27.28), pre-morbidity history of infection and vaccination history(χ²= 9.31), initial ALC (F=12.06) at initial diagnosis were related with the prognosis of children with ITP (all P<0.05). Multivariate analysis showed that the history of infection before onset, the history of vaccination(95%CI: 0.19 to 1.51, P<0.05)and initial ALC(95%CI: -0.64 to -0.23, P<0.001) at the time of initial diagnosis were independent factors affecting the prognosis of children with ITP.The receivers operating characteristic (ROC) was drawn with the development of chronic disease (course >12 months) as state variable and ALC as test variable.The area under curve(AUC) was 0.765(P<0.05), cut-off point was 3.925×10⁹/L, sensitivity was 0.542, and specificity was 0.966. Conclusions Initial ALC can be used as one of the prognosis index of ITP in children, and low ALC is the risk signal for chronic development of ITP in children.

The efficacy of palliative care was definitive with two-dimensional radiotherapy for stage IV non-small cell lung cancer(NSCLC).However,theimpact of radiotherapy on survival was not well indicated,and some study resultsindicating prolonged survival were not accepted. Along with the advancesin three-dimensional radiotherapy (3DRT),wide application of comprehensive treatment,and the understanding of the association between different metastatic status and survival,prospective and retrospective studies have demonstrated that chemotherapy combined with 3DRT for primary tumor is more effective than chemoradiotherapy alone in improving symptoms and prolonging survival,especially for oligometastases of stage IV NSCLC.The dose to primary tumor is closely related to survival,and high-dose radiotherapy may be more likely to prolong survival. Further studies,however,areneeded to take into accountproblems such as thedose,timing,and technical selection of radiotherapy.

Objective To investigate the effects of Danusertib on the changes in Aurora kinase B (Aurora B)/ribosomal protein p70S6 kinase (p70S6K)/ribosomal protein 15 (RPL15) signaling pathway and autophagy in human leukemia cells and its mechanism.Methods Myeloid leukemia cell lines THP-1 and K562 were selected as the research subjects.The experiment was divided into 2 phases.Phase 1:each cell line was treated with the concentration of Danusertib in 0.1 p mol/L,1.0 p mol/L and 5.0 μ mol/L.In control group,2 mL/L dimethyl sulfoxide (DMSO)was given.All the treated cells were cultured for 24 hours.The viability of each cell line was examined by methyhhiazoletrazolium assay and the autophagy was assessed by flow cytometry.In addition,the protein levels of p70S6K,AuroraB,phosphatidylinositol 3-kinase (PI3K),AKT(phosphorylated protein kinase B),mammalian target of rapamycin (mTOR),microtubule-associated protein (LC3),Beclin1 and RPL15 were determined by using Western blot.Part 2:Aurora B and RPL15 were down-regulated in THP-1 and K562 cells,respectively.DMSO was used to dissolve Danusertib(5.0 μ mol/L).The grouping was designed as following:DMSO group (blank control group),Danusertib treated group,empty plasmid group,small interfering RNA(siRNA) group,empty plasmid + Danusertib-treated group and siRNA + Danusertib treated group.The protein levels of Aurora B,p70S6K,RPL15,Beclinl and LC3 were detected by using Western blot.Results (1) Danusertib decreased the viability of THP-1 and K562 cells and the half maximal inhibitory concentration values were 26.9 pmol/L and 30.2 μmol/L for THP-1 and K562 cells,respectively.(2)The protein levels of p-Aurora B/Aurora B,p-p70S6K/p70S6K,RPL15,p-mTOR/mTOR and p-AKT/AKT decreased compared with control cells after being treated with 0.1 μmol/L,1.0 μ mol/L and 5.0 pmol/L of Danusertib in THP-1 and K562 cells,and the differences were statistically significant (all P ＜ 0.05).(3) In THP-1 cells,compared with the empty plasmid group,the protein levels of p70S6K and RPL15 decreased by 22.1％,61.3％ (F =18.1,P =0.001) and 55.4％,56.1％ (F =19.4,P =0.001) in siRNA group and siRNA + Danusertib-treated group after knockdown of Aurora B.In contrast,the protein levels of LC3 increased by 13.6％ and 17.1％ (F =15.4,P =0.001)compared with the empty plasmid group.In addition,the protein levels of Beclin1 and LC3 increased by 39.5％,92.3％ (F=25.2,P=0.001) and 40.2％,58.3％ (F=23.9,P=0.001) in siRNA group and siRNA + Danusertib treated group,compared with the empty plasmid group after down-regulation of RPL15.In K562 cells,compared with the empty plasmid group,the protein levels of p70S6K and RPL15 decreased by 24.2％,62.7％ (F =20.4,P=0.001) and 57.2％,60.1％ (F =23.9,P =0.001) in siRNA group and siRNA + Danusertib treated group after downregulation of Aurora B.But the protein levels of LC3 increased by 17.9％ and 56.7％ (F =20.9,P =0.001)compared with the empty plasmid group.Moreover,the protein levels of Beclin1 and LC3 were increased by 20.6％,98.4％ (F=22.4,P =0.001) and 41.5％,70.1％ (F=26.2,P =0.001) in siRNA group and siRNA + Danusertib treated group,compared with the empty plasmid group after downregulation of RPL15.Conclusion Danusertib can induce autophagy via inhibition of the PI3K/AKT/mTOR signaling pathway and can negatively regulate Aurora B/p70S6K/RPL15 axis in THP-1 and K562 cells.In addition,RPL15 may be a key target of Aurora B/p70S6K/RPL15signaling pathway in the inhibition of tumor proliferation.