BACKGROUND: Cyclooxygenase 2, aggrecanase 1, and insulin-like growth factor 1 are involved in pathological injury of the articular cartilage. OBJECTIVE:To observe the expression of shRNA vectors carrying cyclooxygenase 2, aggrecanase 1 and overexpression vectors carrying insulin-like growth factor 1 in bone marrow mesenchymal stem cels. METHODS:Lentiviral vectors carrying the silencing gene cyclooxygenase 2, aggrecanase 1, the over-expressing gene insulin-like growth factor 1 and binding green fluorescent protein were constructed with recombinant lentiviral technology, and then the recombinant lentiviral vectors were used to transfect passage 3 human bone marrow mesenchymal stem cels culturedin vitro (experimental group). The human bone marrow mesenchymal stem cels transfected with no target gene lentivirals were used as negative control group. The human bone marrow mesenchymal stem cels transfected with no treatment served as blank group. RESULTS AND CONCLUSION:Cyclooxygenase 2 and aggrecanase 1 transfected in human bone marrow mesenchymal stem cels were significantly inhibited at gene and protein levels, while the expression of insulin-like growth factor 1 was increased significantly at gene and protein levels. We confirmed that cyclooxygenase 2 and aggrecanase 1 were successfuly silenced while insulin-like growth factor 1 overexpressed by using lentiviral vectors in human bone marrow mesenchymal stem cels, which brings a new hope for the systemic gene treatment of arthritis.

Objective To establish a high-throughput method that can simultaneously, quickly, and accurately detect main malaria-transmitting Anopheles species and their resistance genes in China, providing a high-throughput monitoring tool for monitoring the main malaria vectors in China after malaria elimination. Methods In different sampling locations, including Tengchong City, Yunnan Province; Wenchang City, Hainan Province; and Donggang City, Liaoning Province, adult specimens of mosquitoes, including Anopheles sinensis, Anopheles minimus, Anopheles dirus, and Anopheles anthropophagus, were collected. Polymerase chain reaction (PCR) technology and Sanger sequencing were employed to detect the ITS2, kdr (L1014), rdl (A296), and ace-1 (G119) genes in individual mosquitoes. For the analysis of mixed samples, an optimized multiplex PCR reaction system, custom-designed dual barcode primers, and next-generation sequencing (NGS) technology were utilized to detect the aforementioned genes. The consistency was assessed using Kappa consistency tests and Chi-square tests for multiple rates. Sensitivity, specificity, and the Youden index were calculated using a four-grid table calculation method. The costs associated with each step of the normal operational process for each method were statistically summarized, and the optimal quantity of mixed samples for detection was determined by a comprehensive approach. Results Conventional PCR amplification of gDNA from 300 mosquitoes resulted in 144 individuals of Anopheles sinensis, 53 individuals of Anopheles dirus, 62 individuals of Anopheles anthropophagus, and 41 individuals of Anopheles minimus, as identified by Sanger sequencing. The mutation frequencies of resistance genes kdr (L1014), rdl (A296), and ace-1 (G119) were found in 73, 27, and 41 specimens, respectively. Using a newly established multiplex PCR reaction system based on custom dual barcode and NGS sequencing technology, samples corresponding to Sanger sequencing were detected under different sample sizes. The two methods showed high consistency in the results (all Kappa>0.900). Multiple comparison tests showed significant differences in the consistencies of the two methods across different sample sizes N (40, 80, 160), N (120, 200, 240, 280), and N (300) (χ2=26.547, P<0.001). The new method demonstrated high sensitivity and specificity across various sample sizes, with the Youden index ranging from highest to lowest as follows: 1 (40, 80, 160)>0.994 (120)>0.990 (280)>0.988 (200)>0.987 (240)>0.985 (300). With an increase in sample size from 40 to 300, the cost per sequencing site for the new method decreased from 20.0 yuan to 8.3 yuan, while the cost per sequencing site for the conventional method decreased from 16.7 yuan to 15.4 yuan. The optimal mixed sample size for the new method was determined to be 280. Conclusion The newly developed multiplex PCR and barcode NGS detection method enables simultaneous screening of four major malaria vector mosquito species and the presence of mutations in the ace-1, kdr, and rdl resistance genes, exhibiting excellent stability, high sensitivity, and specificity. It allows for the efficient analysis of large sample sizes in a single run, offering a cost-effective alternative compared to other types of detection methods.

Aim To investigate the protective effect of cinnamic aldehyde ( CA ) on hormone-induced osteo-clasts proliferation and bone resorption in vitro and its molecular mechanisms. Methods RAW264. 7 cells induced into osteoclast were treated with RANKL and M-CSF and then were divided into control group, dexa-methasone ( DEX ) group and different doses of CA (11. 6, 23. 2, 46. 4 μg·L-1 ) groups. OCs were ob-served after tartrate resistant acid phosphatase( TRAP) staining. The cell proliferation was determined by MTT assay at different time points. The expression levels of TRACP5 b in cell cultured supernatants were measured by ELISA. RT-PCR technique was applied to examine the transcriptional levels of RANK and NFATc1 . Re-sults In MTT assay, the proliferation of osteoclasts stimulated by dexamethasone was promoted seriously compared with negative control group ( P < 0. 05 ) . Meanwhile, DEX could strengthen the content of TRACP5 b and up-regulate the expressions of RANK and NFATc1 mRNA. After administration of CA, the proliferation was inhibited, while the enhanced expres-sion of TRAP5b was reversed,and the over-expressions of RANK and NFATc1 mRNA were obviously down-regulated in a time-and-dose-dependent manner ( P <0. 05 ) . Conclusion The results suggest that CA in-hibits proliferation and bone resorption of osteoclast in-duced by DEX, which may be mediated by down-regu-lation of RANK and NFATc1 mRNA.

Objective To investigate the prevalence of Alzheimer's disease (AD) in first-degree relatives of patients with affective disorders,and to evaluate the risk of AD in first-degree relatives of the patients with affective disorders.Methods Patients with affective disorders meeting DSM-Ⅳ-TR criteria (affective disorders group) and their healthy spouses (conrol group) were recruited in this study (n=109 each).The first-degree relatives inclusion criteria were biological relatives of both probands aged over 55 years.Subjects were investigated by neuropsychological assessment,imaging and clinical examinations,and were diagnosed as AD according to the criteria of the United States of America neuropathy language disorders and stroke research institute and Alzheimer's disease and Related Disorders Association (NINCDS-ADRDA).Results 284 first-degree biological relatives of affective disorders patients and 274 first-degree relatives of control group were selected.There were no statistically significant differences in demographic characteristics of the first-degree relatives between the two groups.The prevalence of Alzheimer's disease in the first-degree relatives had a significant difference between affective disorders group and control group [10.6％ (30/284) vs.4.4％ (12/ 274),x2=7.47,P=0.006].The prevalence of AD in the first-degree relatives was higher in bipolar disorder and depressive disorder patients than in control group [11.2％ (14/125) vs.4.4％(12/274),20.0％ (15/75) vs.4.4％(12/274),x2=6.80,20.56,OR=2.60,4.63,both P＜0.05],while there was no significant difference in the prevalence of AD in the first-degree relatives between mania patients and control group [1.1％ (1/87) vs.4.4％ (12/274),x2 =1.99,P＞0.05].Conclusions There is a high risk for Alzheimer' s disease in first-degree relatives of patients with affective disorders,particularly in first-degree relatives of patients with depression and bipolar disorders.

The purpose of education is to cultivate talents who can master the ability of self-learning lifelong. With the rapid development of multimedia technology, the knowledge carrier represented by micro curriculum plays a very important role in improving students' self-learning ability. In traditional Chinese medicine college, due to the short of time, weak learning foundation, the ability of self-learning is hard to improve in the modern medical courses such as biochemistry. This is not conducive to the cultivation of modern talents of Chinese medicine. In this paper, we chose the biochemistry teaching in TCM college as an example, and discuss how we can make the application of micro courses reasonably in the teaching process. This study was regarded as a starting to improve the students' self-learning ability effectively.

Objective:To investigate the role of dendritic cells(DC) in unexplained recurrent spontaneous abortion(URSA) and to study the effect and molecular mechanism of Traditional Chinese Medicine Shou Tai Decoction ( STD ) on URSA treatment by regulating the function of DC.Methods:30 cases of normal pregnancy women and 30 cases of URSA patients were taken as control group and URSA group respectively.URSA patients were treated with Shou Tai Decoction.Peripheral blood mononuclear cells were taken from both control group and URSA group before and after STD administration.The proportion of CD11c+HLA-DR+cells,CD11c+CD80+cells and CD11c+CD86+cells in peripheral blood were measured by flow cytometry.Moreover,the mRNA expression of HLA-DR,CD80,CD86 and Indoleamine2,3-dioxygenase( IDO) in venous blood were detected by RT-PCR assay.The protein expression of IDO was detected by Western blot.Furthermore, the cytokines, including IL-12p70 and IL-6, in the blood serum were measured by ELISA.Results:Compared with normal pregnancy women,the proportion of CD11c+HLA-DR+,CD11c+CD80+,CD11c+CD86+cells and the mRNA expression of HLA-DR,CD80,CD86 of URSA patients in peripheral blood were both increased significantly(P<0.05), while the mRNA and protein expression of IDO were decreased markedly(P<0.05).Additionally,the level of IL-12p70 and IL-6 in serum of URSA women were significantly increased ( P<0.01 ) .When compared with URSA patients before STD administration, the proportion of CD11c+HLA-DR+,CD11c+CD80+,CD11c+CD86+cells and the mRNA expression of HLA-DR,CD80,CD86 decreased significantly after STD administration ( P<0.05 ) , while the mRNA and protein expression of IDO increased markedly after STD administration(P<0.05).Meanwhile,compared with before STD administration,serum protein level of IL-12p70 and IL-6 of URSA patients decreased significantly after STD treatment ( P<0.01 ) .Conclusion: The changes of proportion and function of DC were involved in URSA.The regulatory effect of STD on DC proportion and function contribute to the treatment of URSA.

Objective To investigate the effects of the point-injection combined with neuromuscular facilitation rehabilitation techniques on post-stroke shoulder-hand syndrome. Methods A treatment group, 36 cases, was treated with the point-injection combined with neural facilitation of rehabilitation techniques. And a control group, 30 cases, was treated with massage therapy. Observed the clinical manifestations and used Fugl-Meyer to assess the joint activities, pain degree and motion function of upper limbs before and after therapy. Results Compared with the control group, the treatment group showed better improvement of joint activity scope and degree, and alleviation of pain (P<0.05). Conclusion Point injection and neuromuscular rehabilitation treatment has a better effect ain treating sequelare of brain stroke and it is worth applying.

Objective:To explore the characteristics of immunological reconstitution of T-cell subsets and the role of Cornus,a Chinese medicinal herb on T lymphocytes in mice after irradiation (IR). Methods:Irradiated model mice were exposed to a single dose of X-ray radiation (2. 6 Gy) with or without Cornus treatments. Bloodroutine was examined before or after irradiation. CD3+,CD4+, CD8+ T cells and Th1, Tc1, Th2, Th17, Treg from spleen or peripheral blood were determined by flow cytometry. Results: Total lymphocytes and CD3+T cells including CD4+T and CD8+T cells,were significantly reduced 3 days after irradiation (P<0. 05). The re-constitution of CD3+ T cells ( especially CD8+T cells ) started from 5 days post irradiation, CD4+T cells increased 8 days after irradiation. However,the production of IFN-γ by Th1 or Tc1 cells were evidently decreased compared with control group even 8 days post irradiation (P<0. 05). In contrast to Th1,Th2/Th17/Treg were observably increased in irradiation group (P<0. 05). But the percentage of Th1 was obviously increased, and Th2, Th17, Treg were markedly decreased in cornus treated mice compared with irradiated mice (P<0. 05). Conclusion: The immunological reconstitution of T lymphocytes started from 5 days after total irradiation with rapid recovery of CD8+ T cells but not CD4+T cells. Cornus effectively improved the imbalance of T cell subsets by promoting the proliferation of Th1 and suppressing Th17 and Treg.

Objective:To demonstrate the relationship between the Th 1/Th2/Th17/Treg paradigm and the bone loss induced by estrogen deficiency and looking for potential target for clinical treatment.Methods:30 BALB/c mice were divided randomly into the normal control group , the sham operation group , and the ovariectomy group.The serum estradiol ( E2 ) was assessed by ELISA.Bone mineral density (BMD) of thigh bone was measured with dual energy X ray absorptiometry.Meanwhile,the T-cell subsets (Th1:CD4+TNFα+, Th2: CD4+IL-4+, Th17: CD4+IL-17 A+, Treg: CD4+CD25+Foxp3+) in spleen lymphocytes were analyzed by flow cytometry.Results:Compared with the normal group and the sham operation group , both E2 and BMD in the ovariectomy group decreased significantly ( P<0.05 ).The percentage of Th 1 and Th17 subset increased while the percentage of Th 2 and Treg decreased significantly in ovariectomy mice compared with sham operation mice.Correlation analysis showed that BMD was positively related to E 2 level and the percentage of Th 2 and Treg subset;however ,BMD was negatively related to the percentage of Th 1 and Th17 subset ( P<0.05 ).Conclusion: Conclusion: T-cell paradigm was involved in the bone loss induced by estrogen deficiency.Modifying T-cell paradigm may become a potential target for reducing bone loss induced by estrogen deficiency .

Objective:To investigate the role of Th1/Th2 shift in unexplained recurrent spontaneous abortion (URSA) and to study the effect and molecular mechanism of Bushen Guchong decoction on URSA treatment by regulating Th 1/Th2 paradigm.Methods:30 cases of URSA patients were given Bushen Guchong decoction administration ; meanwhile,30 cases of normal pregnancy women were taken as control group.Th1/Th2 paradigm in peripheral blood monocytes of URSA patients before and after Bushen Guchong decoction administration was detected by flow cytometry.Moreover,the mRNA expression of the specific transcription factors for Th1 and Th2 subsets -T-bet and GATA-3 were detected by RT-PCR assay.Furthermore,the relationship between the Th1/Th2 paradigm and the uterine bleeding time after Bushen Guchong decoction administration was analyzed by Pearson correlation and re -gression tests.Results: Compared with normal pregnancy women , the proportion of Th1 subset of URSA patients was increased significantly (P<0.05),while the proportion of Th2 subset was decreased markedly (P<0.05).Additionally,the ration of Th1/Th2 subsets was increased in URSA women (P<0.05).After Bushen Guchong decoction treatment ,the proportion of Th1 subsets in URSA patients and T-bet mRNA expression were up-regulated significantly ( P<0.05 ) , whereas the Th2 proportion and GATA-3 mRNA expression were down-regulated obviously (P<0.05).Moreover,the ratio of Th1/Th2 was decreased significantly (P<0.05).Correlation analysis showed that the uterine bleeding time was positively related to both the proportion of Th 1 and the ratio of Th1/Th2, while negatively related to the proportion of Th 2 subsets.Conclusion:Th1/Th2 shift was involved in URSA.Bushen Guchong decoction up-regulated the transcription level of GATA-3 and down-regulated the transcription level of T-bet.Thereby, Bushen Guchong decoction exerted the curative effect on URSA by revising Th 1/Th2 shift.