Construction of a competing endogenous RNA network to identify drug targets against polycystic ovary syndrome.

STUDY QUESTION: Would the construction of a competing endogenous RNA (ceRNA) network help identify new drug targets for the development of potential therapies for polycystic ovary syndrome (PCOS)? SUMMARY ANSWER: Both Food and Drug Administartion (FDA)-approved and candidate drugs could be identified by combining bioinformatics approaches with clinical sample analysis based on our established ceRNA network. WHAT IS KNOWN ALREADY: Thus far, no effective drugs are available for treating PCOS. ceRNAs play crucial roles in multiple diseases, and some of them are in current use as prognostic biomarkers as well as for chemo-response and drug prediction. STUDY DESIGN, SIZE, DURATION: For the bioinformatics part, five microarrays of human granulosa cells were considered eligible after applying strict screening criteria and were used to construct the ceRNA network for target identification. For population-based validation, samples from 24 women with and without PCOS were collected from January 2021 to July 2021. PARTICIPANTS/MATERIALS, SETTING, METHODS: The public data included 27 unaffected women and 25 women with PCOS, according to the Rotterdam criteria proposed in 2003. The limma and RobustRankAggreg R packages were used to identify differentially expressed messenger RNAs and noncoding RNAs. Gene Ontology, Reactome and Kyoto Encyclopedia of Genes and Gemomes (KEGG) enrichment analyses were performed. A ceRNA network was constructed by integrating the differentially expressed genes and target genes. The population-based validation included human luteinized granulosa cell samples from 12 unaffected women and 12 women with PCOS. Quantitative real-time polymerase chain reaction was conducted to detect the levels of mRNAs and microRNAs (miRNAs). Connectivity map and computational model algorithms were implemented to predict therapeutic drugs from the ceRNA network. Additionally, we compared the predicted drugs with known clinical medications in DrugBank. MAIN RESULTS AND THE ROLE OF CHANCE: A set of 10 mRNAs, 11 miRNAs and 53 long non-coding RNAs (lncRNAs) were differentially expressed. Functional enrichment analysis revealed the highest relevance to immune system-related biological processes and signalling pathways, such as cytokine secretion and leucocyte chemotaxis. A ceRNA consisting of two lncRNAs, two miRNAs and five mRNAs was constructed. Through network construction via bioinformatic analysis, we identified some already approved drugs (such as metformin) that could target some molecules in the network as potential drug candidates for PCOS. LARGE SCALE DATA: Public sequencing data were obtained from GSE34526, GSE84376, GSE102293, GSE106724 and GSE114419, which have been deposited in the Gene Expression Omnibus database. LIMITATIONS, REASONS FOR CAUTION: Experiments, such as immunoprecipitation, luciferase reporter assays and animal model studies, are needed to validate the potential targets in the ceRNA network before the identified drug candidates can be tested using cellular and animal model systems. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide new bioinformatic insight into the possible pathogenesis of PCOS from ceRNA network analysis, which has not been previously studied in the human reproductive field. Our study also reveals some potential drug candidates for the future development of possible therapies against PCOS. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by grants from the National Key Research and Development Program of China (2021YFC2700400) and the National Natural Science Foundation of China (82001498). The authors have no conflicts of interest to disclose.

A Pyridazine-Bridged Sandwiched Cluster Incorporating Planar Hexanuclear Cobalt Ring and Bivacant Phosphotungstate.

A planar hexanuclear cobalt ring was clamped by two bivacant α1-[PW10O37](9-) with the assistance of the pyridazine bridges to form a novel sandwiched Co(II)-polyoxometalate cluster compound, [Na(H2O)6][Co3(OH) (pydz)4(H2O)7][Co6(PW10O37)2(pydz)4(H2O)6]·43H2O (1; pydz = pyridazine).This cluster was identified by X-ray single-crystal diffraction, elemental analysis, Fourier transform IR and UV-visible spectroscopies, and cyclic voltammetry (CV). Structural analysis reveals that 1 comprises a hexahydrated sodium, a trinuclear [Co3(OH) (pydz)4(H2O)7](5+) cationic cluster, and an anionic [Co6(PW10O37)2(pydz)4(H2O)6](6-) sandwiched cluster, thus giving an intrinsical intercluster compound. The isolation of such cluster was dependent on the in situ transformation of trivacant [α-P2W15O56](12-) to α1-[PW10O37](9-) under the hydrothermal condition. The CV shows reversible multielectron waves from the redox of W(VI) in 1. Cluster 1 exhibits remarkable electrocatalytic activity toward the reduction of nitrite. Magnetism studies indicated a weak anti-ferromagnetic exchange interaction between Co(II) ions within 1.

Quercetin suppresses HeLa cells by blocking PI3K/Akt pathway.

To explore the effect of quercetin on the proliferation and apoptosis of HeLa cells, HeLa cells were incubated with quercetin at different concentrations. Cell viability was evaluated by MTT assay, cell apoptosis was detected by Annexin-V/PI double labeled cytometry and DNA ladder assay. Cell cycle was flow cytometrically determined and the morphological changes of the cells were observed under a fluorescence microscope after Hoechst 33258 staining and the apoptosis-related proteins in the HeLa cells were assessed by Western blotting. The results showed that quercetin significantly inhibited the growth of HeLa cells and induced obvious apoptosis in vitro in a time- and dose-dependent manner. Moreover, quercetin induced apoptosis of HeLa cells in cell cycle-dependent manner because quercetin could induce arrest of HeLa cells at G0/G1 phase. Quercetin treatment down-regulated the expression of the PI3K and p-Akt. In addition, quercetin could down-regulate expression of bcl-2, up-regulate Bax, but exerted no effect on the overall expression of Akt. We are led to conclude that quercetin induces apoptosis via PI3k/Akt pathways, and quercetin has potential to be used as an anti-tumor agent against human cervix cancer.

Risk factors for recurrence of ovarian endometriomas after surgical excision.

Ovarian endometrioma is a common form of endometriosis, which may cause infertility, dysmenorrhea and pelvic pain in women of reproductive age. Although surgery is the treatment of choice for endometriomas, recurrence poses a formidable frustration. This study investigated potential risk factors of endometriomas recurrence, aiming to better understand its pathogenesis. A total of 307 patients with endometriomas were followed up for an average of 28.6 months and the 1-, 2- and 3-year cumulative recurrence rate was 9.5%, 21.9%, and 29.2%, respectively. Twenty-one potential risk factors for endometriomas recurrence were evaluated using Cox's proportional hazards models. Total revised American Fertility Society (rAFS) score was significantly associated with higher recurrence (OR=1.858, 95% CI=1.122-3.075, P=0.016), as well as younger age at surgery (OR=0.953, 95% CI=0.915-0.992, P=0.020). Semiradical surgical treatment was defined as surgical removal of cyst plus hysterectomy with preservation of bilateral or unilateral ovary, and was a significant factor that was associated with lower recurrence than the conservative surgery (OR=0.318, 95% CI=0.107-0.951, P=0.040). Postoperative pregnancy was favorable factors for disease recurrence (OR=0.217, 95% CI=0.102-0.460, P=0.000). The results suggest that endometrioma recurrence is inversely associated with age at surgery and postoperative pregnancy, and may correlate with total rAFS score and conservative surgery method.

[Pathogenesis and Targeted Treatment Progress of Splenomegaly in Primary Myelofibrosis--Review].

Primary myelofibrosis (PMF) is a myeloproliferative neoplasm with splenomegaly as the major clinical manifestation, which is commonly considered to be linked to splenic extramedullary hematopoiesis. Alteration of CXCL12/CXCR4 pathway can lead to the migration of hematopoietic stem cells and hematopoietic progenitor cells from bone marrow to spleen which results in splenic extramedullary hematopoiesis. In addition, low GATA1 expression and the abnormal secretion of cytokines were found to be significantly associated with splenomegaly. With the application of JAK1/2 inhibitors in clinical, the symptoms of splenomegaly have been significantly improved in PMF patients. This article will review the pathogenesis and targeted treatment progress of splenomegaly in PMF.

Inhibition of Hec1 expression enhances the sensitivity of human ovarian cancer cells to paclitaxel.

AIM: Hec1, a member of the Ndc80 kinetochore complex, is highly expressed in cancers. The aim of this study was to explore the role and mechanism of action of Hec1 with respect to the cytotoxicity of paclitaxel in ovarian cancer. METHODS: Thirty ovarian cancer samples and 6 normal ovarian samples were collected. Hec1 expression in these samples was determined with immunohistochemistry. Ovarian cancer cell lines A2780, OV2008, C13K, SKOV3, and CAOV3 and A2780/Taxol were examined. Cell apoptosis and cell cycle analysis were detected with flow cytometric technique. siRNA was used to delete Hec1 in the cells. The expression of related mRNAs and proteins was measured using RT-PCR and Western blot analysis, respectively. RESULTS: Hec1 expression was significantly higher in ovarian cancer samples than in normal ovarian samples, and was associated with paclitaxel-resistance and poor prognosis. Among the 6 ovarian cancer cell lines examined, Hec1 expression was highest in paclitaxel-resistant A2780/Taxol cells, and lowest in A2780 cells. Depleting Hec1 in A2780/Taxol cells with siRNA decreased the IC50 value of paclitaxel by more than 10-fold (from 590±26.7 to 45.6±19.4 nmol/L). Depleting Hec1 in A2780 cells had no significant effect on the paclitaxel sensitivity. In paclitaxel-treated A2780/Taxol cells, depleting Hec1 significantly increased the cleaved PARP and Bax protein levels, and decreased the Bcl-xL protein level. CONCLUSION: Hec1 overexpression is associated with the progression and poor prognosis of ovarian cancer. Inhibition of Hec1 expression can sensitize ovarian cancer cells to paclitaxel.

[Antitumor and antimetastatic effect of antimicrobial peptide conjugated with tumor homing peptide TMTP1 on the transplanted prostate cancer and gastric cancer in nude mice].

OBJECTIVE: Due to their lower risk for induction of resistance, antimicrobial peptides with selective anticancer effect could be developed into a new generation of anticancer drugs. We conjugated an antimicrobial peptide with tumor-targeting peptides (TMTP1) to explore whether it has inhibiting effect on the progression and metastasis of transplanted prostate cancer and gastric cancer in nude mice. METHODS: Subcutaneously transplanted human prostate cancer and orthotopically transplanted human gastric cancer in nude mice were prepared. 50 µmol/L PBS (control group), 50 µmol/L TMTP1 (TMTP1 group) or 50 µmol/L TMTP1-GG-D(KLAKLAK)(2) (treatment group) were injected i.p. to the three groups of nude mice, respectively. The binding ability of the novel fusion polypeptide TMTP1-GG-D(KLAKLAK)(2) to the tumors and its antitumor effect were assessed by measurement of tumor volume, histopathological examination of the tumor tissues, testing apoptosis index of tumor cells with TUNEL staining, and survival curve plotting of the mice. RESULTS: The median survival time of subcutaneous prostate cancer-bearing mice was 50 days in the control group, 55 days in the TMTP1 group, and 70 days in the TMTP1-GG-D(KLAKLAK)(2) group (P < 0.05). The median survival time of the subcutaneous gastric cancer-bearing mice was 25 days in the control group, 30 days in the TMTP1 group, and 45 days in the TMTP1-GG-D(KLAKLAK)(2) group (P < 0.01). The tumor volume in the subcutaneous prostate cancer-bearing mice was (2.5 ± 0.3)cm(3) in the control group, (1.8 ± 0.2) cm(3) in the TMTP1 group, and (0.3 ± 0.1)cm(3) in the TMTP1-GG-D(KLAKLAK)(2) group (P < 0.01). The tumor volume of the subcutaneous gastric cancer-bearing mice was (3.8 ± 0.4) cm(3) in the control group, (3.2 ± 0.2)cm(3) in the TMTP1 group, and (0.4 ± 0.1) cm(3) in the TMTP1-GG-D(KLAKLAK)(2) group (P < 0.01). Large tumors were observed in the stomach of the orthotopic gastric cancer-bearing mice of the control and TMTP1 groups. The tumor volume of the TMTP1-GG-D(KLAKLAK)(2) group was obviously reduced. White metastases in the liver, spleen and abdominal wall were observed in the control and TMTP1 groups (P < 0.01). TUNEL staining revealed that the apoptosis index of the control group was (31.9 ± 1.5)%, TMTP1 group (37.2 ± 2.3)% and TMTP1-GG-D(KLAKLAK)(2) group (69.7 ± 2.1)% (P < 0.01). CONCLUSIONS: The results of our study demonstrate that the novel fusion peptide of antimicriobial peptide conjugated with TMTP1 can effectively inhibit tumor progression and metastasis, therefore, is promising to be a novel effective anticancer drug.

HPV16 E5 peptide vaccine in treatment of cervical cancer in vitro and in vivo.

Human papillomavirus (HPV)-induced cervical cancer is the second most common cancer among women worldwide. Despite the encouraging development of the preventive vaccine for HPV, a vaccine for both prevention and therapy or pre-cancerous lesions remains in high priority. Thus far, most of the HPV therapeutic vaccines are focused on HPV E6 and E7 oncogene. However these vaccines could not completely eradicate the lesions. Recently, HPV E5, which is considered as an oncogene, is getting more and more attention. In this study, we predicted the epitopes of HPV16 E5 by bioinformatics as candidate peptide, then, evaluated the efficacy and chose an effective one to do the further test. To evaluate the effect of vaccine, rTC-1 (TC-1 cells infected by rAAV-HPV16E5) served as cell tumor model and rTC-1 loading mice as an ectopic tumor model. We prepared vaccine by muscle injection. The vaccine effects were determined by evaluating the function of tumor-specific T cells by cell proliferation assay and ELISPOT, calculating the tumor volume in mice and estimating the survival time of mice. Our in vitro and in vivo studies revealed that injection of E5 peptide+CpG resulted in strong cell-mediated immunity (CMI) and protected mice from tumor growth, meanwhile, prolonged the survival time after tumor cell loading. This study provides new insights into HPV16 E5 as a possible target on the therapeutic strategies about cervical cancer.

[Association of FOXP3 gene polymorphism in Chinese women with endometriosis].

OBJECTIVE: To assess the association between single nucleotide polymorphisms (SNPs) of forkhead box P3 gene (FOXP3) and endometriosis in Chinese Han women from central China. METHODS: MassARRAY IPLEX and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) technique was used to determine the genotypes of FOXP3 gene in 314 patients with endometriosis and 358 healthy controls. RESULTS: Genotypes of C/T polymorphism for the rs2280883 locus, A/C for the rs3761548 locus, and C/T for the rs3761549 locus were determined. No significant difference was detected in distribution of genotypes CC, CT and TT (P=0.770, OR=0.960; P=0.923, OR=1.013) and frequencies of C and T alleles (P=0.772, OR=0.960; P=0.925, OR=1.013) for rs2280883 and rs3761549 between the two groups. And no significant difference was detected in distribution of genotypes AA, AC and CC (P=0.762, OR=0.958) and frequencies of A and C alleles (P=0.715, OR=0.950) for rs3761548 was detected between the two groups. Based on r-AFS classification, the patients were divided into two groups (respectively with I-II stage and III-IV stage endometriosis). Again, no significant difference was detected in distribution of genotypes CC, CT and TT (P=0.454, OR=1.198, P=0.526, OR=0.909; P=0.220, OR=0.750, P=0.548, OR=1.094) and frequencies of C and T alleles (P=0.473, OR=1.215, P=0.532, OR=0.912; P=0.204, OR=0.737, P=0.558, OR=1.089) for rs22080883 and rs3761549 loci between the two patient groups. No association was found between distribution of genotypes AA, AC and CC (P=0.431, OR=1.211; P=0.508, OR=0.905) and frequencies of A and C alleles (P=0.417, OR=1.226; P=0.516, OR=0.908) for rs3761548 locus between the two patient groups. CONCLUSION: Our study has failed to found any association between FOXP3 gene polymorphisms rs2280883, rs3761548 and rs3761549 with endometriosis in Chinese Han patients.

[Protective effects of caloric restriction on ovarian function].

OBJECTIVE: To study the protective effects on ovarian function by caloric restriction (CR) and its mechanism. METHODS: Thirty female C57BL/6 mice of 8 weeks old were randomly divided into two groups, including ad libitum (AL) group and caloric restriction (CR) group. The general situation and ovarian function of those mice were compared and evaluated.Ovarian follicles were counted by hematoxylin-eosin staining. Anti-Miillerian Hormone(AMH) mRNA expression of the ovary were detected by using real-time PCR. The concentrations of serum estradiol, progesterone of the mice were measured by ELISA. And the fertility of mice by mating trials were evaluated, SIRT3, Hypoxia inducible factor 1α (HIF-1α) and catalase (CAT) mRNA expression of the mice ovaries were detected by Real-Time PCR. RESULTS: The total follicles were 546 in CR mice and 286 in AL mice. The proportion of primordial follicles were 38.6% (211/546) in ovaries of CR mice and 29.4% (84/286) in ovaries of AL mice, which reached statistical difference. The proportion of atretic follicles 5.3% (29/546) in ovaries of CR mice, compared with 16.8% (48/286) in AL mice, was significantly decreased (P < 0.05). The AMH mRNA expression in CR mice ovaries was 3.37 times of that of AL mice (P < 0.05). The serum concentration of estradiol in CR mice was up to (5.3 ± 1.6) pmol/L, which was much higher than (3.6 ± 1.6) pmol/L in AL mice. While, the progesterone concentration of (0.4 ± 0.3) nmol/L in CR mice was lower than (1.4 ± 0.8) nmol/L in AL mice (P < 0.05).Fertility and survival of offsprings were both improved in CR mice. The expression level of SIRT3 mRNA in CR mice ovary was 1.39 times, CAT was 1.55 times and HIF-1α was 0.31 times of those in AL mice (P < 0.05). CONCLUSIONS: Caloric restriction can delay the ovary aging process through reduce follicle depletion by suppressing follicle recruitment and ovulation. The function of ovarian reserve and reproductive endocrine was effectively protected. Caloric restriction can reduce the incidence of follicular atresia, its mechanism might be associated with anti-oxidative stress.

Efficacy of Bispecific Antibody Targeting EpCAM and CD3 for Immunotherapy in Ovarian Cancer Ascites: An Experimental Study.

OBJECTIVE: This study aimed to explore the value of M701, targeting epithelial cell adhesion molecule (EpCAM) and CD3, in the immunotherapy of ovarian cancer ascites by the in vitro assay. METHODS: The expression of EpCAM in ovarian cancer tissues was analyzed by databases. The EpCAM expression and immune cell infiltration in different foci of ovarian cancer were detected by 8-channel flow cytometry. The toxic effect of M701 on OVCAR3 was tested using the in vitro cytotoxicity assay. The 3D cell culture and drug intervention experiments were performed to evaluate the therapeutic effect of M701 in ovarian cancer specimens. Flow cytometry was used to examine the effect of M701 on the binding of immune cells to tumor cells and the activation capacity of T cells. RESULTS: The results of the bioinformatic analysis showed that the expression of EpCAM in ovarian cancer tissue was significantly higher than that in normal ovarian tissue. The 8-channel flow cytometry of clinical samples showed that the EpCAM expression and lymphocyte infiltration were significantly heterogeneous among ovarian cancer patients and lesions at different sites. The in vitro experiment results showed that M701 had a significant killing effect on OVCAR3 cells. M701 also obviously killed primary tumor cells derived from some patients with ovarian cancer ascites. M701 could mediate the binding of CD3+ T cells to EpCAM+ tumor cells and induce T cell activation in a dose-dependent manner. CONCLUSION: M701 showed significant inhibitory activity on tumor cells derived from ovarian cancer ascites, which had a promising application in immunotherapy for patients with ovarian cancer ascites.

Extracellular matrix-derived scaffolds in constructing artificial ovaries for ovarian failure: a systematic methodological review.

STUDY QUESTION: What is the current state-of-the-art methodology assessing decellularized extracellular matrix (dECM)-based artificial ovaries for treating ovarian failure? SUMMARY ANSWER: Preclinical studies have demonstrated that decellularized scaffolds support the growth of ovarian somatic cells and follicles both in vitro and in vivo. WHAT IS KNOWN ALREADY: Artificial ovaries are a promising approach for rescuing ovarian function. Decellularization has been applied in bioengineering female reproductive tract tissues. However, decellularization targeting the ovary lacks a comprehensive and in-depth understanding. STUDY DESIGN SIZE DURATION: PubMed, Embase, Web of Science, and the Cochrane Central Register of Controlled Trials were searched from inception until 20 October 2022 to systematically review all studies in which artificial ovaries were constructed using decellularized extracellular matrix scaffolds. The review was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocol. PARTICIPANTS/MATERIALS SETTING METHODS: Two authors selected studies independently based on the eligibility criteria. Studies were included if decellularized scaffolds, regardless of their species origin, were seeded with ovarian cells or follicles. Review articles and meeting papers were removed from the search results, as were articles without decellularized scaffolds or recellularization or decellularization protocols, or control groups or ovarian cells. MAIN RESULTS AND THE ROLE OF CHANCE: The search returned a total of 754 publications, and 12 papers were eligible for final analysis. The papers were published between 2015 and 2022 and were most frequently reported as coming from Iran. Detailed information on the decellularization procedure, evaluation method, and preclinical study design was extracted. In particular, we concentrated on the type and duration of detergent reagent, DNA and extracellular matrix detection methods, and the main findings on ovarian function. Decellularized tissues derived from humans and experimental animals were reported. Scaffolds loaded with ovarian cells have produced estrogen and progesterone, though with high variability, and have supported the growth of various follicles. Serious complications have not been reported. LIMITATIONS REASONS FOR CAUTION: A meta-analysis could not be performed. Therefore, only data pooling was conducted. Additionally, the quality of some studies was limited mainly due to incomplete description of methods, which impeded specific data extraction and quality analysis. Several studies that used dECM scaffolds were performed or authored by the same research group with a few modifications, which might have biased our evaluation. WIDER IMPLICATIONS OF THE FINDINGS: Overall, the decellularization-based artificial ovary is a promising but experimental choice for substituting insufficient ovaries. A generic and comparable standard should be established for the decellularization protocols, quality implementation, and cytotoxicity controls. Currently, decellularized materials are far from being clinically applicable to artificial ovaries. STUDY FUNDING/COMPETING INTERESTS: This study was funded by the National Natural Science Foundation of China (Nos. 82001498 and 81701438). The authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: This systematic review is registered with the International Prospective Register of Systematic Reviews (PROSPERO, ID CRD42022338449).

Efficacy and Safety of the Chinese Herbal Compound TJAOA101 in Treating Diminished Ovarian Reserve: A Protocol for Multicenter, Prospective, and Pre-Post Study.

OBJECTIVE: Diminished ovarian reserve (DOR) can lead to early menopause, poor fecundity, and an increased risk of disorders such as osteoporosis, cardiovascular disease, and cognitive impairment, seriously affecting the physical and mental health of women. There is still no safe and effective strategy or method to combat DOR. We have developed a novel Chinese herbal formula, Tongji anti-ovarian aging 101 (TJAOA101), to treat DOR. However, its safety and efficacy need to be further validated. METHODS: In this prospective and pre-post clinical trial, 100 eligible patients aged 18-45 diagnosed with DOR will be recruited. All participants receive TJAOA101 twice a day for 3 months. Then, comparisons before and after treatment will be analyzed, and the outcomes, including anti-mullerian hormone (AMH) and follicle-stimulating hormone (FSH) levels and the antral follicle count (AFC), the recovery rate of menopause, and the Kupperman index (KMI), will be assessed at baseline, every month during medication (the intervention period), and 1, 3 months after medication (the follow-up period). Assessments for adverse events will be performed during the intervention and follow-up periods. CONCLUSION: A multicenter, prospective study will be conducted to further confirm the safety and efficacy of TJAOA101 in treating DOR and to provide new therapeutic strategies for improving the quality of life in DOR patients.

[Preparation of HPV18 E7 peptide plus CpG vaccine and its immunologic effects in vitro].

OBJECTIVE: To screen and prepare the vaccine of human papillomavirus (HPV) 18 E7 peptide target at human leukocyte antigen (HLA)-A2 plus CpG through SYFPEITHI. METHODS: (1) The SYFPEITHI database was employed for predicting and screening of HPV18 E7 HLA-A2 restricted T cell epitopes.(2) The peripheral blood and tumor tissue sample of HLA-A2 positive and HPV18 positive/negative patients were collected and randomly divided into 7 groups, i.e. E7PA + CpG, E7PB + CpG, E7PC + CpG, E7PD + CpG, CpG, IR-T + CpG and control groups respectively. T cell proliferation was detected by thiazolyl blue tetrazolium bromide (MTT) assay at different timepoints. Lactate dehydrogenase delivery method (LDH) was used to test the cytolytic t lymphocyte (CTL) activity of peripheral blood mononuclear cell (PBMC) in different ratios of effect and target (E:T). And the level of activity T cells was evaluated by interferon gamma (IFN-γ)-related enzyme-linked immuno-spot assay (ELISPOT). RESULTS: (1) Four peptides named E7PA, E7PB, E7PC and E7PD were obtained separately with high levels of affinity and specificity. (2) During continuous observations after vaccination, the E7PA + CpG group had the most pronounced proliferation rate. When E:T = 100:1, the E7PA + CpG group had more powerful CTL effect than the control group with statistic significance (P < 0.00). E:T was concentration-dependent. Except for IR-T + CpG, all other groups had great difference than control group with statistic significance (P < 0.05) but no significant difference between the groups. The levels of IFN-γ spot-forming T cells were higher in the E7PA + CpG group than the control group with statistic significance (P < 0.01). In terms of specificity, E7PA + CpG in the HPV18 positive group could induce the proliferation of IFN-γ-secreting T cells. And there was statistical difference with the control group (P < 0.05). CONCLUSION: Screening the HPV18 E7 peptide target at HLA-A2 plus CpG as the candidate targets by SYFPEITHI may active specific immunological cellular responses to HPV18 positive disease.

[Expression and clinical significance of Sema4C in esophageal cancer, gastric cancer and rectal cancer].

OBJECTIVE: To explore the expression and clinical significance of signal protein Sema4C in esophageal cancer, gastric cancer and rectal cancer. METHODS: Fifty esophageal cancer, 75 gastric cancer, 50 rectal cancer and 20 corresponding normal mucous membrane specimens, collected during the period of January 2008 to December 2010, were detected with streptavidin-peroxidase immunohistochemistry to detect the expression levels of Sema4C. And the relationships of the Sema4C expression with clinicopathological data was analyzed. RESULTS: The expression levels of Sema4C in three kinds of cancers were significantly higher than the corresponding normal mucous membranes (80.0% (n = 40) vs 20.0% (n = 4), 77.3% (n = 58) vs 25.0% (n = 5), 80.0% (n = 40) vs 15.0% (n = 3), all P = 0.000). Furthermore, the percentage of Sema4C positive cells was significantly higher in carcinoma nests of tumors with lymphatic metastasis than those without (90.3% (n = 28) vs 63.2% (n = 12), 85.0% (n = 51) vs 46.7% (n = 7), 92.0% (n = 23) vs 68.0% (n = 17), P = 0.049, 0.005, 0.034). However, no significant correlations were found between the Sema4C expression with gender, age, location of tumors, types of cancer cells, cell differentiation, tumor size, depth of invasion or tumor stage (all P > 0.05). CONCLUSION: There is a high expression of Sema4C in esophageal cancer, gastric cancer and rectal cancer. And it is strongly correlated with lymphatic metastasis. Thus Sema4C may play critical roles in the invasion and lymphatic metastasis of esophageal cancer, gastric cancer and rectal cancer.

[Research Advance of the Mechanisms, Clinical Characteristics and Treatment Strategy of Coagulation Dysfunction in Hemophagocytic Lymphohistiocytosis --Review].

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening disease caused by uncontrolled proliferation of activated macrophage, and secreting high amounts of inflammatory cytokines which lead to multi-organ dysfunction syndrome. HLH patients often show different clinical characteristics during the disease was progressed, in which coagulopathy were the most common, including thrombocytopenia and hypofibrinogenemia, those are the major cause of death in patients, and the clinicians should increase awareness of the mechanisms, clinical characteristics, prognosis and treatment. In this review, the above problems are briefly summarized, to deepen understanding of the HLH related coagulation dysfunctions, and early identification and treatment to reduce mortality, so as to provide more opportunities for HLH patients to recieve subsequent treatment.

Construction of Artificial Ovaries with Decellularized Porcine Scaffold and Its Elicited Immune Response after Xenotransplantation in Mice.

Substitution by artificial ovary is a promising approach to restore ovarian function, and a decellularized extracellular matrix can be used as a supporting scaffold. However, biomimetic ovary fabrication and immunogenicity requires more investigation. In this study, we proposed an effective decellularization protocol to prepare ovarian scaffolds, which were characterized by few nuclear substances and which retained the extracellular matrix proteins. The ovarian tissue shape and 3-dimensional structure were well-preserved after decellularization. Electron micrographs demonstrated that the extracellular matrix fibers in the decellularized group had similar porosity and structure to those of native ovaries. Semi-quantification analysis confirmed that the amount of extracellular matrix proteins was reduced, but the collagen fiber length, width, and straightness did not change significantly. Granulosa cells were attached and penetrated into the decellularized scaffold and exhibited high proliferative activity with no visible apoptotic cells on day 15. Follicle growth was compromised on day 7. The implanted artificial ovaries did not restore endocrine function in ovariectomized mice. The grafts were infiltrated with immune cells within 3 days, which damaged the artificial ovary morphology. The findings suggest that immune rejection plays an important role when using artificial ovaries.

[Immune Thrombocytopenia in Patients with Chronic Lymphocytic Leukemia: Pathological Mechanism and Treatment Progress---Review].

Chronic lymphocytic leukemia (CLL) patients usually show immune dysfunction, which often leads to autoimmune hemocytopenia. Immune thrombocytopenia (ITP) is one of the common complications. The pathogenesis of CLL-related ITP is complex and has not been fully elucidated. At present, the researches mainly focus on humoral immunity, cellular immunity and innate immune disorders. Recent studies suggest that genomic abnormalities and microRNAs are also involved in CLL-related ITP. Traditional ITP standard therapy has a poor effect on CLL-related ITP. Chemotherapy or monoclonal antibody therapy against the primary pathogenesis of CLL can effectively treat thrombocytopenia, and the emergence of new targeted drugs also provides new treatment options for the disease. In this paper, the progresses of CLL-related ITP pathogenesis, prognosis and treatment in recent years are reviewed.

Clinicopathologic Characteristics and Survival of Patients with Rare Malignant Ovarian Yolk Sac Tumors: A Population-based Analysis.

Yolk sac tumors (YSTs) are rare malignant germ cell tumors that usually affect young females. To date, there have been few studies on YSTs. We evaluated the relationship between clinicopathologic characteristics of patients with ovarian YSTs and disease outcome based on Surveillance, Epidemiology, and End Results data. The Kaplan-Meier method and log-rank test were used to evaluate differences in survival rates. Data for 269 patients were analyzed. The incidence of YSTs among ovarian germ cell tumors (OGCTs) cases was 0.4%; median patient age was 22.0 years, and most tumors were unilateral. Patients presented with distant metastasis (37.5%), localized disease (49.1%), and regional spread (8.9%). American Joint Committee on Cancer stage was available for 13 patients (stage IA, n=2; stage IC, n=1; stage IIIA, n=1; stage IIIB, n=3; stage IIIC, n=2; and stage IV, n=4). Survival rates at 1, 3, and 5 years were 91.0%, 84.0%, and 83.2%, respectively, for overall survival (OS) and 92.0%, 85.4%, and 84.5%, respectively, for disease-specific survival (DSS). The 5-year OS and DSS of patients with ovary tumors were 91.5% and 92.9%, respectively, compared to 74.8% and 77.2%, respectively, for those with extra-ovarian spread (P<.001 for both OS and DSS). Age >50 years was associated with shorter OS and DSS (both P<0.001), whereas no associatios of OS and DSS were observed with pathologic grade (P=0.49 for OS and 0.52 for DSS). In summary, YSTs are typically unilateral, of a high grade, and localized to the ovary; extra-ovarian spread has a poor outcome, and postmenopausal women have worse prognosis than premenopausal women.

[Effect of HPV16 peptide vaccine in combination with paclitaxel-cisplatin chemotherapy on cervical cancer in vitro and in vivo].

OBJECTIVE: to the explore the effect of Human papillomavirus (HPV) 16 peptide vaccine in combination with paclitaxel-cisplatin (TP) chemotherapy on cervical cancer in vitro and in vivo. METHODS: (1) the major histocompatibility complex (MHC) class I restricted T cell epitopes were studied by bioinformatics for transporter associated with antigen processing (TAP). Their effects were compared and E7Pa had the most dramatic effect. (2) In vivo, the C57BL/6 mice were divided equally into 6 groups randomly after loading with TC-1 cells (HPV 16 positive tumor cells from C57BL/6 mouse), named as E7Pa + CpG + TP, E7Pa + CpG, CpG + TP, TP and CpG group as experiment groups and control (blank injection with physiological saline). The tumor volumes were measured regularly by tumor growth curve to compare the therapeutic effects in different groups; the related cell factors in murine peripheral blood were evaluated by enzyme-linked immunosorbent assay (ELISA); the TUNEL test kit was used to explore cellular apoptosis in tumor tissue; the survival curve was drawn from the TC-1 cell loading to natural death; safety was tested by pathological test and leucocyte count. RESULTS: at day 60 of tumor growth, the tumor volume of immunotherapy plus TP chemotherapy group was (0.013 ± 0.010) cm(3) versus the control (1.900 ± 0.075) cm(3) with a great significant deviation (P < 0.01). Meanwhile, the volumes were E7Pa + CpG group (0.340 ± 0.038) cm(3), TP + CpG group (0.650 ± 0.029) cm(3), TP group (1.100 ± 0.052) cm(3) and that of CpG group was (0.890 ± 0.047) cm(3) separately. And these groups had significant difference with the controls (P < 0.05). The average survival time of different groups were E7Pa + CpG + TP group (108.50 ± 8.97) d, E7Pa + CpG group (100.02 ± 2.27) d, CpG + TP group (79.63 ± 4.05) d, TP group (73.24 ± 3.11) d, CpG group (68.63 ± 1.38) d and controls (52.37 ± 2.47) d. And the difference between the E7Pa + CpG + TP and E7Pa + CpG groups had great significance with the controls (P < 0.01). Furthermore, the immune system was effectively stimulated for suppressing tumor growth in the immunotherapy group while cell apoptosis was significantly induced in the chemotherapy group. The combination of immunotherapy and chemotherapy was significantly efficacious than either of them alone. And it could thoroughly stimulate the immune effects and enhance the anti-tumor function of chemotherapeutical drugs. In safety test, there was no significant difference among all groups. CONCLUSION: the HPV16 peptide vaccine in combination with TP chemotherapy can treat the HPV16 E7 positive tumor effectively in experiment.