The clinicopathologic significance of the loss of BAF250a (ARID1A) expression in endometrial carcinoma.

OBJECTIVE: AT-rich interactive domain 1A (ARID1A) is a tumor suppressor gene that encodes the BAF250a protein. Recent studies have shown the loss of ARID1A expression in several types of tumors. We aimed to investigate the clinical and pathologic role of BAF250a in endometrial carcinoma. METHODS: We examined the expression of BAF250a and its correlation with the expression of p53, estrogen receptor, progesterone receptor, glucocorticoid receptor, hypoxiainduciblefactor-1α, and vascular endothelial growth factor in normal and various malignant endometrial tissues. RESULTS: The expression of BAF250 was significantly down-regulated in endometrial carcinoma when compared with normal endometrial tissues. The loss of BAF250a expression was found in 25% of endometrial carcinoma samples but not in normal endometrial tissues, complex endometrial hyperplasia, and atypical endometrial hyperplasia samples. Subtypes of endometrial carcinoma, especially uterine endometrioid carcinoma and uterine clear cell carcinoma, had higher frequency of loss of BAF250a expression. In addition, the expression of BAF250a was positively correlated with estrogen receptor and negatively correlated with p53 in poorly differentiated endometrial adenocarcinoma. Moreover, the expression of BAF250a was significantly associated with the differentiation status of endometrial carcinoma but not associated with clinical stage, the depth of myometrial invasion, lymph node metastasis, and overall survival of patients with endometrial carcinoma. CONCLUSIONS: Our data showed that loss of BAF250a is frequently found in high-grade endometrioid and clear cell carcinomas but not in other types of endometrial carcinoma. The loss of BAF250a expression does not have prognostic value for endometrial carcinoma.

Establishing a new animal model for hepadnaviral infection: susceptibility of Chinese Marmota-species to woodchuck hepatitis virus infection.

Hepatitis B virus infection (HBV) is a major medical problem in China. The lack of a suitable infection model in China is recognized as an obstacle for research on HBV in China. Chinese Marmota-species is phylogenetically closely related to Marmota monax, thus, it might be suitable to serve as an animal model for HBV infection. Therefore, we attempted to prove the claim about the existence of woodchuck hepatitis virus (WHV)-like viruses in Chinese Marmota-species and to determine the susceptibility of these species to experimental WHV infection. In the present study, 653 sera from three Chinese Marmota-species, Marmota himalayana, Marmota baibacina and Marmota bobak, were screened for WHV-like viruses by serological and molecular assays. The susceptibility to WHV of three species was investigated by experimental infection and monitored by testing of anti-WHc and WHsAg by ELISA, detection of WHV DNA by PCR, and detection of WHV replication intermediates and antigens in liver samples. No evidence for the existence of a genetically closely related virus to WHV in three Chinese Marmota-species was found by serological assays and PCR. M. himalayana was susceptible to WHV infection as inoculated animals became positive for anti-WHc, WHsAg and WHV DNA. Further, WHV replication intermediates and proteins were detected in liver samples. In contrast, M. baibacina remained negative for tested virological parameters. M. bobak species showed a limited susceptibility to WHV. Our data do not support early reports about WHV-like viruses in China. M. himalayana is suitable for the establishment of a model for hepadnaviral infection.

Creation of a planned or central-clefted puncture combined with a second puncture during vertebroplasty to treat osteoporotic vertebral compression fractures with large clefts.

BACKGROUND: Cemented vertebrae frequently re-fracture after vertebroplasty to treat osteoporotic vertebral compression fractures (OVCFs) with large clefts. We compared the efficacy of planned and central-clefted puncture, both followed by a second puncture, as treatments for OVCFs with large clefts. METHODS: We retrospectively studied 38 patients. 18 of whom underwent planned puncture (group A) and 20 central-clefted puncture (group B). A second puncture was performed when the initially injected cement was restricted to the cleft. We recorded a visual analog scale (VAS) pain scores, vertebral kyphotic angles (KAs), and compression ratios (CRs) preoperatively and at 2 days and 6 months postoperatively. We recorded the cement dispersion patterns and complications. RESULTS: Second punctures succeeded in 15/18 and 7/20 patients of groups A and B, respectively. At 2 days postoperatively, the VAS score, KA, and CR were significantly better than the preoperative values (P < 0.01); no significant difference was found between the two groups (P > 0.05). At the 6-month follow-up, all scores were poorer than at 2 days postoperatively (all P < 0.05), significantly more so in group B than group A (P < 0.05). Significant differences in terms of the cement dispersion patterns, and the cemented vertebral re-fracture and cement leakage rates, were observed between the two groups (all P < 0.05). CONCLUSION: The two-puncture techniques were initially effective when treating large-clefted OVCFs. However, compared to the central-clefted puncture, the planned puncture improved the success rate of the second puncture, allowed better cement dispersion, and reduced the incidence of vertebral re-fracture during follow-up.

[Inhibitory effects of enhanced expression of CD40L in ovarian cancer OVHM cells on the liver metastases in mice].

OBJECTIVE: To examine the in vivo anti-metastatic effect of enhanced expression of CD40L cDNA in murine ovarian cancer OVHM cells (CD40L-OVHM) injected into the spleen on liver metastasis in mice. METHODS: OVHM cells were inoculated into the spleen of 6 to 8 week-old female B6C3F1 (C57BL/6N x C3H/He) mice. The established liver metastasis was identified by histopathology (HE staining). OVHM cells, DNA-pMKITneo-OVHM cells or CD40L-OVHM cells were inoculated into the spleen of female B6C3F1 mice and the expressions of CD11c in splenic cells were detected by flow cytometry. The specific cytotoxicity of splenic cells was detected by MTT assay, and the serum cytokines of IFN-gamma, TNF-alpha, IL-12, IL-4 and IL-10 of the mice were measured by enzyme linked immunoabsorbent assay. The liver metastases and the survival time of the mice were also recorded. RESULTS: The mouse models with liver metastasis by injecting tumor cells into the spleen of mice were established. The expression of CD11c and the specific killing rate in CD40L-OVHM cells group was significantly higher than that in the OVHM cells group and DNA-pMKITneo-OVHM cells group. The expressions of IFN-gamma, TNF-alpha and IL-12 in the CD40L-OVHM cells group were much more increased than OVHM cells group and DNA-pMKITneo-OVHM cells group, but the expressions of IL-4 and IL-10 in the CD40L-OVHM cells group were decreased significantly (p < 0.05). The average weights of livers and spleens of mice in CD40L-OVHM cells group were significantly lower than those of DNA-pMKITneo-OVHM cells group and OVHM cells group. The survival time of mice in CD40L-OVHM cells group was also significantly longer than that in the OVHM cells group and DNA-pMKITneo-OVHM cells group (P < 0.05). CONCLUSION: The data directly demonstrate that the expression of CD40L in ovarian cancer cells (CD40L-OVHM) can enhance the proliferation and differentiation of dendritic cells in the spleen and induce specific cytotoxic effect of T cells in the spleen, and may regulate the immune function of peripheral blood cells and the immune balance between Th1 cells and Th2 cells, which maybe the possible mechanism induced by CD40L in mice inhibiting the development of liver metastasis.

[Enhanced expression of CD40L cDNA on ovarian cancer cell line OVHM induces the secretion of Th1 cytokines from dendritic cells].

OBJECTIVE: To examine whether the enhanced expression of CD40L cDNA on murine ovarian cancer (OVHM) cells could induce the secretion of Th1 cytokines from dendritic cells (DC). METHODS: OVHM cells were transfected with the full-length mouse CD40L cDNA by lipofectamine 2000 and then G418 resistant cells as positive cells were selected. They were examined for their expression of CD40L with flow cytometry. Bone marrow cells were firstly depleted of erythrocytes, macrophages, T and B cells with PE-conjugated magnetic beads, and then cultured in 10% FCS RPMI 1640 medium supplemented with recombinant mouse GM-CSF and IL-4 for 10 days. PKH67-labeled tumor cells were cultured with DC, and then the stained cells were analyzed for the expression of MHC-I, MHC-II, CD80, CD86, CCR7 in DC with flow cytometry. The expression of p40, p19, p35, p28, EBI3 subunits, IL-18, IFN-gamma, Mig gene in cocultured DC-tumor cells were detected by RT-PCR. RESULTS: The CD40L cDNA was successfully transfected into OVHM cells. Bone marrow-derived DCs, when cultured with CD40L/OVHM, formed clusters with the tumors and showed an upregulated expression of MHC- I, MHC-II, CD80, CD86, CCR7. Expression of the IL-12, IL-23, IL-27, IL-18, interferon-gamma (IFN-gamma) and Mig (monokine induced by IFN-gamma) genes was induced in the DCs that were cultured with CD40L/OVHM but not with OVHM cells. CONCLUSION: These data directly showed that the expression of CD40L on ovarian cancer cells facilitates the interaction between DCs and tumors, enhances the maturation of DCs, induces secretion of Th1 cytokines, especially for IL-12, IL-23 and IL-27, which maybe one of the possible antitumor mechanism for CD40L-transfected ovarian cancer cell line.

[The growth inhibition effects of TSLC1 gene on human hepatocyte carcinoma cell line HepG2].

OBJECTIVES: To study the effects of tumor suppressor in lung cancer-1 (TSLC1) on human hepatocyte carcinoma cell line HepG2. METHODS: A full length of TSLC1 cDNA was amplified from RNA of normal human liver cells by RT-PCR, and it was cloned into a pCI-neo expression vector and transfected into human hepatocellular carcinoma cell line HepG2. The HepG2 cells transfected with this plasmid (experimental group) and those treated with pCI-neo vector (control group) and without any treatment (blank group) were compared. Cell morphology was studied microscopically and cell growth was analyzed with MTT assay. FACSort flow cytometry analysis was performed to assess the cell cycle distribution and apoptosis. RESULTS: A stable cell line expressing TSLC1 protein was successfully established. Morphologically, cells of the experimental group were tightly aggregated when compared with those of the control and blank groups. The growth of TSLC1-transfected cells was significantly suppressed in vitro compared with those of the control and blank groups. The amount of G0-G1 cells was 63.66%+/-3.83% (P less than 0.01) in the experimental group, while those of the control and blank groups were 47.45%+/-0.91% and 54.47%+/-0.96% respectively. The amount of S phase cells in the experimental group, 22.90%+/-6.04%, was significantly lower (P less than 0.05) than that of the control group (36.58%+/-0.61%) and the blank group (33.61%+/-2.99%), which suggested a G0-G1 cell cycle arresting. The number of cells in early and late phase apoptosis (17.09%+/-0.20% and 16.11%+/-0.40% respectively) were significantly higher than those of the control and blank groups (P less than 0.01). CONCLUSIONS: TSLC1 strongly inhibits the growth of HepG2 cells in vitro and induces apoptosis of the cells, suggesting that TSLC1 may have a tumor suppressor function in HCC.

[Antigenicity of major hydrophilic region II of hepatitis B virus surface antigen].

OBJECTIVE: To study whether the substitutions at the major hydrophilic region II (MHRII) of hepatitis B surface antigen (HBsAg) will impair the antigenicity of HBsAg. METHODS: Four recombinant plasmids expressing mutant HBsAg (mtHBsAg) P1120T, C121S, K122I and T123N were constructed. HepG2 cells were transfected with the four plasmids and a plasmid expressing G145R HBsAg. The immunoreactivity of the cells expressing mtHBsAg with P1120T, C121S, K122I, T123N and G145R were detected by immunofluorescence (IF) staining and ELISA with 4 antibodies and 7 HBsAg diagnostic kits respectively. RESULTS: mtHBsAg with P120T was recognized by mAb1 and mAb2. mtHBsAg with C121S and K122I was not recognized by any mAbs. mtHBsAg with T123N in lysates was recognized by mAb2, but not recognized in the supernatants. CONCLUSION: Substitutions at amino acid positions 120-123 of HBsAg strongly impaired the antigenicity of HBsAg, a fact that was not appreciated previously.

Inhibition of hepatitis B virus replication by APOBEC3G in vitro and in vivo.

AIM: To investigate the effect of APOBEC3G mediated antiviral activity against hepatitis B virus (HBV) in cell cultures and replication competent HBV vector-based mouse model. METHODS: The mammalian hepatoma cells Huh7 and HepG2 were cotransfected with various amounts of CMV-driven expression vector encoding APOBEC3G and replication competent 1.3 fold over-length HBV. Levels of HBsAg and HBeAg in the media of the transfected cells were determined by ELISA. The expression of HBcAg in transfected cells was detected by western blot. HBV DNA and RNA from intracellular core particles were examined by Northern and Southern blot analyses. To assess activity of the APOBEC3G in vivo, an HBV vector-based model was used in which APOBEC3G and the HBV vector were co-delivered via high-volume tail vein injection. Levels of HBsAg and HBV DNA in the sera of mice as well as HBV core-associated RNA in the liver of mice were determined by ELISA and quantitative PCR analysis respectively. RESULTS: There was a dose dependent decrease in the levels of intracellular core-associated HBV DNA and extracellular production of HBsAg and HBeAg. The levels of intracellular core-associated viral RNA also decreased, but the expression of HBcAg in transfected cells showed almost no change. Consistent with in vitro results, levels of HBsAg in the sera of mice were dramatically decreased. More than 1.5 log10 decrease in levels of serum HBV DNA and liver HBV RNA were observed in the APOBEC3G-treated groups compared with the control groups. CONCLUSION: These findings indicate that APOBEC3G could suppress HBV replication and antigen expression both in vivo and in vitro, promising an advance in treatment of HBV infection.

[Inhibition of hepatitis B and duck hepatitis B virus replication by APOBEC3G].

OBJECTIVE: To investigate the effect of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G) mediated antiviral activity against hepatitis B virus (HBV) and duck hepatitis B virus (DHBV). METHODS: Total RNA was extracted from peripheral blood mononuclear cells (PBMCs), RT-PCR product was cloned into the EcoR I/Hind III restriction sites of the CMV-driven expression vector fused with a hemagglutinin fusion epitope tag at its carboxyl terminal. Replication competent 1.3 fold over-length HBV was constructed with full-length HBV of ayw subtype. The mammalian hepatoma cell HepG2 was cotransfected with the replication competent 1.3 fold over-length HBV and various amounts of CMV-driven expression vector encoding APOBEC3G-HA. Levels of HBsAg and HBeAg in the media of the transfected cells were determined by ELISA, HBV DNA. RNA from intracellular core particles was examined using Northern and Southern blot analyses. Chicken hepatoma cell LMH was cotransfected with head-to-tail dimer of an EcoR I monomer of DHBV and various amounts of CMV-driven expression vector encoding APOBEC3G-HA. DHBV DNA from intracellular core particles was examined using Southern blot analysis. RESULTS: CMV-driven expression vector encoding APOBEC3G-HA and replication competent 1.3 fold over-length HBV were constructed. There was a dose dependent decrease in the levels of intracellular core-associated viral (HBV and DHBV) DNA and extracellular production of HBsAg and HBeAg. Levels of intracellular core-associated viral RNA were also decreased, but the expression of HBcAg remained almost unchanged. CONCLUSION: APOBEC3G suppresses HBV and DHBV replication and also suppresses HBsAg and HBeAg expression.

Antitumor effects and mechanisms of dendritic cells stimulated by sCD40L on ovarian cancer cells in vitro.

OBJECTIVE: This study aimed to examine the expression of immune suppression factors and the mechanisms of antitumor effects of cord blood dendritic cells (DCs) stimulated by soluble cluster of differentiation 40 ligand (sCD40L) and cytokines in vitro in ovarian cancer patients. METHODS: The expression levels of interleukin (IL)-10 and transforming growth factor (TGF)-ß messenger RNA in peripheral blood were detected by reverse transcription polymerase chain reaction; expression levels of CD80 and CD86 in DCs stimulated by sCD40L were detected using flow cytometry and confocal laser scanning microscopy. RESULTS: Expression levels of IL-10 and TGF-ß genes in the peripheral blood of ovarian cancer patients were significantly increased compared with patients with benign ovarian tumors (P < 0.05). The expression levels of CD80 and CD86 in DCs cultured in the granulocyte-macrophage colony-stimulating factor + IL-4 + stem cell factor + Flt-3 ligand + sCD40L group were significantly increased compared with those in the control group, as assessed by flow cytometry and confocal laser scanning microscopy (P < 0.05). CONCLUSION: A variety of cytokines in combination with sCD40L can promote the proliferation of cord blood-derived DCs and induce their maturation as well as stimulating a specific antitumor response.

Antidiabetic effects of Artemisia sphaerocephala Krasch. gum, a novel food additive in China, on streptozotocin-induced type 2 diabetic rats.

AIM OF THE STUDY: Since ancient times, practicians of traditional Chinese medicine have discovered that Artemisia sphaerocephala Krasch. (Asteraceae) seed powder was useful for the treatment of diabetes. Artemisia sphaerocephala Krasch. gum (ASK gum), which is extracted from seed powder of the plant, is a novel food additive favored by the food industry in China. The objective of this study was to determine the antidiabetic function of ASK gum on type 2 diabetes. MATERIALS AND METHODS: Type 2 diabetic rat model was induced with high fat diet and low dose of streptozotocin (STZ). The effects of ASK gum on hyperglycemia, hyperlipemia, insulin resistance, and liver fat accumulation in type 2 diabetic rats were evaluated. The results were compared to those of normal rats and diabetic rats treated with metformin. RESULTS: The addition of ASK gum to the rats' food supply significantly lowered fasting blood glucose, glycated serum protein, serum cholesterol, and serum triglyceride in type 2 diabetic rats, and significantly elevated liver glucokinase, liver glycogen, and serum high density protein cholesterol in the diabetic rats. ASK gum significantly reduced insulin resistance and liver fat accumulation of type 2 diabetes. CONCLUSION: Artemisia sphaerocephala Krasch. gum can alleviate hyperglycemia, hyperlipemia and insulin resistance of type 2 diabetes.

[Establishment of an in vivo model for duck hepatitis B virus infection using Hubei duckling].

OBJECTIVE: To develop a standard duck hepatitis B virus (DHBV) animal model using a local Hubei species of duck, Ma Ya, and use it as an in vivo experimental system to study antiviral strategies against hepatitis B. METHODS: Two-day-old Ma Ya ducklings were experimentally infected via intraperitoneal injection with the DHBV inocula which was collected from the transfected culture supernatant of 1.5-fold-overlength genome recombinant plasmid. Blood samples were taken twice or thrice a week during post-inoculation for 50 days. Viremia was quantified by serum real-time PCR to show the peak. Antiviral treatment of the DHBV-infected ducklings was started 3 d post-inoculation. The animals received oral administration of lamivudine (3TC) at a dose of 25 mg/kg/d for 5 d, followed by a maintenance therapy thrice weekly for 3 more weeks. Serum was quantified to show the viremia peak and liver biopsy specimens were analysed by Southern blotting and in-situ hybridization at the end of antiviral drug treatment. RESULTS: The experimental infection rate of 2-day-old ducklings was 87.5%. Viremia started to be detectable on day 7 and reached a peak on day 11 post-inoculation, followed by a decrease and fluctuations. Four weeks of oral administration of 3TC led to a significant decrease in viremia peak during. This effect was not sustained, as a rebound in viremia was observed after drug withdrawal. Similarly, the analysis of liver biopsies at the end of 3TC treatment showed a marked decrease in DHBV DNA. However, after drug withdrawal a rebound of intrahepatic DHBV DNA was observed in duck livers. CONCLUSION: The Hubei duck model with experimental DHBV infection of transfected supernatant is more suitable for the hepadnavirus biologic research due to its stability and practicability.

[Biological characteristics of an established model of ovarian cancer in mice and its homologous cell lines].

BACKGROUND & OBJECTIVE: There are no specific methods for early diagnosis of ovarian cancer, recurrence prevention and drug-resistance. The experimental mouse model of ovarian cancer could help to reveal the biological and genetic features of ovarian cancer, and provide rational basis for further intervention strategy. This study was to establish a model of ovarian cancer in mice and homologous cell line, and analyze its biological characteristics. METHODS: Ovarian cancer was developed in 8-week-old female F1 (C57BL/6N x C3H/He) mice by a single whole-body neutron irradiation of 2.7 Gy from a (252)Cf source. A metastatic cell line was established through serial subcutaneous transplantation of the primary tumor for 11 generations, and then tumor cells were transferred to in vitro cultivation. These cells were cloned for more than 6 months. The biological characteristics of the tumors and the homologous cell line were determined by cellular and molecular biological techniques. RESULTS: The grafted tumors in mice were successively passaged for 11 generations with a successful inoculation rate of 96% during 23 months. A tumor cell line OV99 isolated from the grafted tumors was established after 6 months and grew steadily. Morphologic characters and ultrastructures of OV99 cells were accorded with those of ovarian cancer epithelia. The chromosomal analysis of OV99 cells revealed aneuploid pattern of 76 chromosomes. Flow cytometry (FCM) and reverse transcription-polymerase chain reaction (RT-PCR) showed same features between OV99 cells and positive control ovarian cancer cell line OVHM, including distribution of cell cycle, rapid growth rate and the expression of P21, P185, P53, proliferating nuclear cell antigen (PCNA) and Cyclin D proteins, and MAGE-1 and MAGE-3 mRNA. CONCLUSION: Establishment of the ovarian carcinoma animal model in mice and OV99, a cell line owns biologic characteristics of ovarian cancer cells, provides experimental materials for further investigation of ovarian carcinoma.

[Study of cytokines secreted by tumor-draining lymph node cells from lung cancer patients induced with different stimulators].

AIM: To investigate the influence of different stimulators on cytokines secretion of tumor-draining lymph node (TDLN) cells and study antitumor effects and mechanism of TDLN cells. METHODS: IL-12 p70, IFN-gamma, TNF-alpha and NO which were secreted by TDLN cells induced by different stimulators (IL-2 group, IL-2+autologous tumor antigen group, IL-2+GM-CSF+IL-4+autologous tumor antigen group and IL-2+GM-CSF+IL-4+LPS group) were detected by ELISA and Griess after TDLN cells were stimulated for 7, 14 and 21 d. RESULTS: The rate of CD83(+) cells of TDLN cells induced by IL-2+GM-CSF+IL-4+autologous tumor antigen and IL-2+GM-CSF+IL-4+LPS was significantly increased. The TDLN cells in the two groups secreted much more IL-12 p70, IFN-gamma and TNF-alpha than IL-2 group and IL-2+autologous tumor antigen group. The TDLN cells stimulated by IL-2+GM-CSF+IL-4+LPS secreted significantly more NO than the other three groups. TDLN cells in each group secreted IL-12 p70, IFN-gamma and NO at the highest level after TDLN cells were stimulated for 14 d. CONCLUSION: Different stimulators can make TDLN cells produce different number of CD83(+) cells (DC), which gives TDLN cells different antitumor activity.

[Biological and antitumor effects of CD40L on ovarian cancer cell line OVHM].

BACKGROUND & OBJECTIVE: Accumulative evidences have indicated the importance of CD40-CD40L receptor-ligand pair in the initiation and regulation of human immune response. Whether triggering CD40-CD40L signaling pathway could induce antitumor effects in ovarian cancer cells is still unclear. This study was to investigate the biological and antitumor effects of CD40L on ovarian cancer cell line OVHM (CD40-positive). METHODS: CD40L cDNA was transfected into OVHM cells by lipofectamine 2000. Immunofluorescent technique and flow cytometry were used to evaluate the expression of CD40, CD40L, MHC-I, MHC-II, CD80, CD86. Cell proliferation was detected by MTT assay. CD40L/OVHM and OVHM cells were inoculated hypodermically into C57BL/6N x C3H/He F1 (B6C3F1) mice and BALB/c nude mice simultaneously. Tumor volume was measured. The production of IFN-gamma secreted by the spleen cells from CD40L/OVHM and OVHM cell-inoculated mice co-cultured with 20 Gy-irridiated OVHM cells were measured with enzyme-linked immunosorbent assay (ELISA). RESULTS: Expression of CD40L in OVHM/CD40L cells was significantly higher than that in OVHM cells (P<0.05). Moreover, the expression of co-stimulatory molecules CD80 and CD86 of CD40L/OVHM was increased (P<0.05). The proliferation rate of parental and CD40L-expressed OVHM cells were not different. Although, delayed tumor growth was observed when CD40L-expressing OVHM cells and OVHM cells were injected simultaneously and contralaterally, the tumor growth of OVHM and CD40L/OVHM in BALB/c nude mice was not different. The amounts of IFN-gamma secreted by the co-culture of irradiated OVHM cells with spleen cells from mice injected with CD40L/OVHM were much higher than those from OVHM/parental cell inoculated and naive mice. CONCLUSION: The forced expression of CD40L cDNA in OVHM cells can increase the immunogenicity, and thus develop antitumor effects against OVHM-incubated mice. In vivo immunization of immunocompetent mice with CD40L/OVHM improves the generation of cytotoxic lymphocytes against parent ovarian cancer cells.