Objective To study the function of the nano-molecular polyamide-amine (PAMAM) as microRNA(miR) carrier targeting gastric adenocarcinoma, and the foundation of developing an efficient delivery of small molecule drugs tar-geting gastric cancer thereof. Methods The folic acid (FA)/PAMAM comoles compound was prepared by dialysis method. After transfection of miR-7 or liposomes into SGC-7901 cell line, fluorescence microscope was used to detect the gene trans-fect efficiency. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the miR-7 level. The immu-nocytochemistry assay was used to test the protein expressions of epidermal growth factor receptor (EGFR), protein kinase B (PKB) and proliferating cell nuclearantigen (PCNA). The transwell system was utilized to explore the migration ability of tu-mor cells. Results Compared with liposme, FA/PAMAN complex compound can significantly improve the level of miR-7 in SGC-7901 cells,reduce the protein levels of EGFR, PKB and PCNA in SGC-7901 cells, and also reduce the percentage of cancer cell migration (P<0.05). Conclusion PAMAM can effectively transfect miR into gastric cancer cells, which is expected to become an efficient delivery of small molecule drugs.

Objective　To evaluate the therapeutic efficiency of reversed lamellar keratoplasty for bullous keratopathy, Terrien's marginal degeneration and ocular chemical burns.Methods　Twenty eyes of patients with bullous keratopathy, twelve eyes with Terrien's marginal degeneration, ten eyes with chemical burns were recruited in this study.Diseased tissues were removed by means of dissection.Reversed lamellae corneal tissues which had been prepared by cryopreservation were thawed and rinsed. Then the prepared posterior corneal lamellae was turned over, endothelium facing upwards. The diameter of the graft should be 0.5～1.0mm larger than that of the recipient bed. The reversed donor posterior lamellae was sutured into the host keratectomy bed with 10-0 nylon sutures continuously or interruptedly.Results　Allograft reaction episode occurred only one (chemical burn) of the total 42 eyes enrolled.The grafts of the rest 41 eyes maintained clear or semitransparent for 1～5 years. Graft edema could be observed within 2～4 weeks postoperatively. Postoperative interface separation was not found. For all 20 patients with bullous keratopathy, the symptoms of pain disappeared. The visual acuity improved 2～3 lines of Snellen among eleven of twelve patients with Terrien's marginal degeneration. The visual acuity improved 2～3 lines of Snellen with four patients with chemical burns.Conclusion　The technique of reversed lamellar keratoplasty can be applied successfully for tectonic reconstruction of corneal surface such as bullous keratopathy, Terrien' s marginal degeneration and also chemical burns.The effects of therapy was satisfactory.

Objective To explore the regulation mechanism for miR - 125a - 5P in epidermal growth factor receptor(EGFR)signaling pathway in medulloblastoma. Methods The potential targets of miR - 125a - 5P in the EGFR signaling pathway were predicted by TargetScan and Sanger software,there were 3 groups:control group,non -sense group and miR - 125a - 5P group. Their relationship,between miR - 125a - 5P and cyclin - dependent kinase in-hibitor 2B( CDKN2B),E2F transcription factor 3( E2F3),mitogen - activated protein kinase 14( MAPK14)and growth factor receptor - bound protein 10(GRB10),were tested by luciferase experiments. After miR - 125a - 5P oligo-nucleotide was transfected to D341 cells,miR - 125a - 5P level was detected by reverse transcription polymerase chain reaction. Then the thiazolyl blue tetrazolium bromide assay was used to draw the cell growth curves,and Transwell assay was used to detect cell migration ability. The expression levels of GRB10,EGFR,phosphatidylinositol 3 - kinase(PI3K) and Ras were tested by Western blot method. Results The results of luciferase experimental results showed that GRB10 was the only target gene of miR - 125a - 5P. After miR - 125a - 5P being transfected,the D341 cell prolifera-tion obviously declined markedly. Compared with control group[(38. 16 ± 7. 47)% ]and the non - sense group [(36. 79 ± 8. 94)% ],cell migration rate in the miR - 125a - 5P group was lowest[(13. 59 ± 4. 41)% ],and there was a significant difference among 3 groups(χ2 = 11. 495,P < 0. 05);in the miR - 125a - 5P group,the expression level of EGFR increased 1. 67 times,GRB10,PI3K and Ras levels were reduced to 23% ,61% and 42% . Conclusion miR - 125a - 5P can inhibit tumor growth by silenced GRB10 expression targeting EGFR downstream signaling pathways in medulloblastoma.

Objective To analysis the malignant performance characteristics of tumor stem cell-like side popula-tion cells in patients with cervical cancer. Methods The cervical cancer cells were obtained from surgical resection tumor tissue. The tumor stem cell-like side population cells were isolated by flow cytometry. The cell growth curve was drawn by MTT assay. The invasion ability of tumor cells was compared by transwell assay. The clonogenic capacity was detected by clone formation in soft agar. The expression level of ABCG2 protein, a drug-resistant gene, was detected by immunofluores-cence method. Finally, these cells were transplated into the subcutaneous of de thymus mice. The rate of tumor formation was compared between groups. Results The results from flow cytometry assay showed the percentage of cervical cancer stem cell-like side population cells was 1.39%. Compared with the non-side population cells, the side population cells grow quickly, showed the enhanced invasion ability and colony forming ability. There was more high expression level in ABCG 2 protein of side population cells. The tumor form rate was 100%(10/10) in the side population cells and the non-side popula-tion cells was 20%(2/10). Conclusion The cervical cancer stem cell-like side population cells have more malignant perfor-mance characteristics than that of non-side population cells, which maybe a core target for cancer gene therapy in the future.

Objective To study the functional roles of 1,25(OH)2D3 in osteogenic differentiation of the dental papilla stem cells. Methods The dental papilla stem cells were isolated and cultured in medium supplemented with different con-centrations of 1,25(OH)2D3 (1, 10 and 100 nmol/L). MTT assay was used to detect the cell growth, and flow cytometry was used to detect the cell cycle. Western blot assay was used to detect protein expression levels of receptor activator of nuclear factor-κB ligand (RANKL), osteoprotegerin (OPG) and vitamin D receptor (VDR). After siRNA silencing VDR expression, protein levels of RANKL and OPG were detected. Results MTT and flow cytometry results showed that there were no sig-nificant differences in the cell proliferation between different concentrations of 1,25(OH)2D3 (1, 10 and 100 nmol/L) and con-trol groups (P>0.05). Western blot results showed that there were protein expressions of VDR, RANKL and OPG in control group. The protein expressions of VDR, RANKL and OPG were increased after adding 1,25(OH)2D3, in which the upward trend was the most significant in VDR. After VDR expression was silenced by siRNA, the protein expression levels of VDR, RANKL and OPG were decreased. Conclusion 1,25(OH) 2D3 affects the osteoblast differentiation process of the dental pa-pilla stem cells by adjusting the VDR expression.

Objective To study the effect of carbonated drinks on primary and permanent teeth replacement in Chil-dren. Method Dog tooth enamel samples were soaked in coca-cola, sprite and pure soda, and the calcium, phosphorus lev-el were analysed. Dental papilla stem cells were separated and cultured in the conditioned medium by adding three drinks. PCR and western blot were used to detect mRNA and protein levels of activator of nuclear factor-k B receptor ligand (RANKL), osteoprotegerin (OPG) and vitamin D receptor (VDR) , then the possible role of each gene and interactions rela-tionship were analyzed. Results Compared with saline, coca-cola and sprite showed their significantly decalcification and dephosphorization role, while plain soda water showed calcium and phosphorus protective effect. These three drinks had no effect on mRNA and protein levels of RANKL gene (P>0.05). Coca-cola and sprite can reduce OPG mRNA and protein lev-els, and at the same time increase transcription and expression of the VDR gene. Plain soda water has no effect on the OPG gene manifestation, but can significantly reduce the transcription and translation level of the VDR gene. Conclusion Car-bonated drinks may affect the dental health of the children's primary and permanent teeth replacement by regulating bone re-lated gene expression and vitamin D receptor family.

With the development of bio-technological drugs, drug immunogenicity evaluation has become key factor of clarifying safety and efficacy of these drugs. It has become the focus to establish a stable and reliable evaluation system. Due to the advantages such as continuous real-time monitoring, surface plasmon resonance (SPR) technology has been widely used in bio-technological drugs immunogenicity assessments. Our study applied this technology to detect anti-drug antibody (ADA) of a recombinant human anti-rabies monoclonal antibody NM57 in the sera of 48 volunteers admitted in phase I clinical trials. This method could satisfy the basic requirements of detection of ADA.

Objective:To observe the ferroptosis in hippocampal neurons of rats with multiple cerebral concussion (MCC) .Methods:Ninety clean grade male Wistar rats with a body mass of (250±10) g were randomly divided into control group (12 rats) and model group (78 rats) according to the random number table method. The rat of MCC model was prepared by hitting the frontotemporal lobe of rats with free fall method once a day for 3 consecutive days.Then the MCC model rats were randomly divided into 12 h, 24 h, 48 h, 72 h, 7 d and 14 d groups with 12 rats in each group. The balance beam experiment was used to detect the motor coordination function of the rats. The levels of interleukin-β(IL-1β), interleukin-6(IL-6), serum glutathione (GSH), neuron-specific enolase (NSE) in serum of rats were detected by ELISA.The content of iron ion in hippocampus was detected by colorimetry. The mRNA and protein levels of glutathione peroxidase 4(GPX4), ferritin heavy (FTH) and ferritin light(FTL) in the hippocampus were detected by RT-PCR and Western blot. Prussian blue staining was used to observe the iron deposition in brain tissue.The ultrastructural changes of hippocampal neurons and mitochondria were observed by transmission electron microscope. SPSS 24.0 statistical software was used for one-way ANOVA among groups, and LSD test was used for multiple pairwise comparison.Results:In the balance beam experiment, the passing time and motor coordination score of rats in each group were significantly different ( F=30.08, 60.34, both P<0.05). The passing time and motor coordination score of rats in the 48 h group ((87.00±4.74) s, (4.75±0.43)) were significantly higher than those in the control group ((35.13±6.99) s, (0.75±0.23)) (both P<0.05). There was significant difference in the total iron ion content, Fe 2+ content and Fe 3+ content in hippocampus of rats in each group ( F=25.20, 94.42, 40.25, all P<0.05), and the content of Fe 2+ in hippocampus of 48 h group was significantly higher than that of the control group ((10.17±0.05) ng/μL, (8.65±0.01) ng/μL)( P<0.05). In the results of RT-PCR and Western blot, the mRNA and protein levels of GPX4, FTH and FTL in hippocampus of each group were significantly different ( F=37.94, 82.09, 49.01, 71.63, 28.94, 15.78, all P<0.05). The mRNA level and protein level of GPX4 ((1.09±0.01), (0.23±0.01) )and FTL ((1.60±0.03), (0.64±0.02)) in 24 h group were significantly higher than those of the control group (GPX4: (1.00±0.02), (0.17±0.01)), FTL: ((1.00±0.04), (0.32±0.01))(all P<0.05). The mRNA level and protein level of FTH ((0.24±0.03), (0.07±0.01)) in 24 h group were significantly lower than those of the control group((1.00±0.01), (0.67±0.03))(both P<0.05). The results of electron microscope showed that the hippocampal neuronal cells of the model rats were reduced, the nucleolus was broken and the nuclear membrane was shrunk in varying degrees, the mitochondria were swollen and deformed and there were vacuoles, and the cristae in the mitochondria decreased or disappeared. Conclusion:The levels of inflammatory reaction and oxidative stress in the multiple cerebral concussion model rats increase, and the hippocampal neurons show the characteristics of ferroptosis, especially at 24 h and 48 h.

Objective The aim of this study was to investigate the anti-tumor effect on sequential injection of heterogeneic lymphocyte(HL)and autogeneic lymphocyte(AL). Methods The HL was prepared by using CC3HF1 mice as feeders. CB6F1 mice were used as recipients,and Hepa1-6 cells were inoculated into the recepients′groin subcutis. A cryoprecipitate was extracted from mouse plasma by freeze-thaw method to prepare fibrin Glue(FG);FG was combined with HL or AL to be FG-HL or FG-AL. The experimental treatment consisted of two stages. At first stage(15 d),FG-HL were injected on the surface of the tumor-bearing tissue of the recipients as the experimental group,and FG-phosphate buffer saline(FG-PBS)were injected on the surface of the tumor-bearing tissue of the rest recipients as the control group. The immunological factors such as tumor cell killing rate of the spleen lym-phocytes and numbers of lymphocytes,CD8 +T and NK in the two groups were detected,respectively. At later stage(10 d),a part of mice were randomly selected from the experimental and control groups,and the lymphocytes( AL) were used to form FG-AL,which were injected on the surface of tumor-bearing tissues in the rest of mice. Tumors in mice of the two groups were compared for tumor volume and tumor inhibition rate. Results The tumor cell killing rate of AL in the experimental group(26. 70 ± 7. 22) was signifi-cantly higher than that in the control group(5. 70 ± 2. 68)(P<0. 01). Numbers of mouse spleen lymphocytes,CD8 +T cells and NK cells were significantly higher than the corresponding values of the control group(P<0. 05). After the two-stage treatment,the aver-age tumor volume of the experimental group[(1.20 ±0.33)cm3]was significantly smaller than that of control group[(2.05 ±0.37) cm3](P<0. 01). The tumor inhibition rate in the experiment group was 41. 5% when compared to the control group. Conclusion Local injections of FG-HL followed by FG-AL can significantly inhibit the growth of transplanted tumor in mice;it is expected to become an anti-tumor biological therapy.

【Objective】 To analyze the clinical and pathological characteristics of mucinous tubular and spindle cell carcinoma （MTSCC） in the kidney so as to evaluate the value of radiotherapy in local treatment of refractory MTSCC. 【Methods】 A case of MTSCC with recurrent recurrence and metastasis was reported， and the existing literature on MTSCC was reviewed and analyzed to explore the value of radiotherapy in the treatment of MTSCC. 【Results】 After three operations， postoperative pathology of the patient showed mucinous tubular and spindle cell carcinoma of the kidney. Radiotherapy was performed in the first recurrence area， and long-term follow-up in the tumor bed area showed no recurrence. 【Conclusion】 MTSCC， a rare renal epithelial tumor， has the low-grade malignance in most cases， and rarely recurs and has metastasis. This patient had repeated recurrence and metastasis before surgery， but after tumor bed radiotherapy after the first recurrence no lesion was found， suggesting that radiotherapy plays a certain role in preventing local recurrence. It is necessary to further explore the value of radiotherapy in the treatment of MTSCC.