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Objective:To evaluate the effects of high mobility group protein box 1 (HMGB1) on clinical prognosis of esophagus squamous cell carcinoma (ESCC) patients treated with chemoradiotherapy and the radiosensitivity of xenograft in nude mice.Methods:A total of 90 endoscopic biopsy specimens were obtained from ESCC patients treated with chemoradiotherapy. The expression level of HMGB1 was determined by immunohistochemical staining. High expression level was defined when staining was observed on ≥50% of the tumor cells. All patients were divided into the high expression group ( n=48) and low expression group ( n=42), and their survival information was retrospectively analyzed. Cell transfection was performed with the plasmid carrying human HMGB1-shRNA to knockdown HMGB1 expression in ECA109 cells and xenograft mouse models were established. The tumor volume and mass were calculated after irradiation with a dose of 15 Gy. The cell apoptosis in xenograft tissues were detected. Survival analysis was performed using Kaplan-Meier method. Univariate prognostic analysis was conducted by log-rank test. Intergroup comparison was performed by analysis of variance (ANOVA). Results:The expression level of HMGB1 was significantly associated with gross tumor volume, longest diameter of tumor, T staging and distant metastasis ( χ2=9.663, 5.625, 4.068, 7.146, all P<0.05). In the low expression group, the overall survival (OS) ( χ2=4.826, P=0.028), progression-free survival (PFS) ( χ2=4.390, P=0.036) were longer compared with that in the high expression group. Further analysis of HMGB1-high expression patients showed that the radiation dose and the combination of chemoradiotherapy did not significantly affect the OS or PFS of ESCC patients. We observed that knockdown of HMGB1 slowed the growth rate of xenograft, decreased the tumor volume and increased the apoptosis rate after irradiation. Conclusions:ESCC patients with high expression level of HMGB1 obtain poor prognosis after chemoradiotherapy, which can be enhanced by increasing the sensitivity to radiotherapy and chemotherapy. HMGB1 knockdown can effectively increase the radiosensitivity of xenograft in ESCC nude mice.
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Objective:To evaluate the effect of compound UC2288 on the radiosensitivity of CNE-2R cell line and nude mouse transplanted tumor.Methods:The UC2288 concentration was referenced to previous experimental results (IC 50=12.20 μmol/L). The effect of UC2288 combined with 2, 4, 6, 8 Gy X-ray irradiation on the radiosensitivity of CNE-2R cell line was detected by clone formation experiment. The effect of UC2288 combined with 2, 4, 6, 8 Gy X-ray irradiation on the proliferation of CNE-2R cell line was determined by CCK8 assay. The nude mouse model of transplanted tumor was constructed with CNE-2R cell line. The radiosensitivity of transplanted tumor of UC2288 combined with 2 Gy/fraction X-ray irradiation for three consecutive days was evaluated. Results:The experimental concentration of UC2288 was 8 μmol/L. The clonality of CNE-2R cell line was reduced under UC2288 combined with X-ray 2, 4, 6, and 8 Gy irradiation, andthe radiosensitizationratio was 1.60. The proliferation of CNE-2R cell line was significantly decreased under UC2288 combined with X-ray 2, 4, 6, and 8 Gy irradiation. UC2288 inhibited the growth of transplanted tumor in nude mice, and the inhibitory effect was strengthened with the extension of observation time, and the most obvious effect was observed at 16 d. ( P<0.01). Theradiosensitizationratio was 4.33. The proliferation of CNE-2R cell line was decreased under UC2288 combined with X-ray irradiation. Conclusion:UC2288 can increase the radiosensitivity of nasopharyngeal carcinoma radioresistant cell line CNE-2R.
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Objective To explore the relationship between miR-26a and metadherin (MTDH), and to verify the relationship between miR-26a and MTDH in vivo in nude mice. Methods Immunohistochemical SP staining method was used to detect the expression of MTDH and in situ hybridization was used to detect the expression of miR-26a in 86 cases of esophageal cancer, and the correlation between the expressions was analyzed. The bioinformatics prediction Targetscan Human 7. 2 software could predicte the binding fragment of MTDH on the miR-26a sequence, and the luciferase report experiment was used to verify the targeted regulatory relationship between MTDH and miR-26a. Nude mice were injected esophageal cancer cell lines subcutaneously which were lentiviral interferenced and overexpressed miR-26a to observe the fonnation of tumors. The tumors from nude mice were made into paraffin, and each was detected. The expression of MTDH in miR-26a in the tumor groups was detected by immunohistochemical staining and in situ hybridization and the relationship was analyzed. Results The expression of miR-26a in esophageal cancer tissues was significantly lower than that in paired nonnal esophageal tissues, and the expression of MTDH in esophageal cancer tissues was significantly higher than that in paired normal esophageal tissues. The expression of miR-26a was related to the patient' s pathological grade (P<0. 05), N stage(P<0. 05), and tumor volume (P<0. 01). The expression of MTDH in esophageal cancer was related to the N stage (P<0. 05) and the degree of differentiation (P<0. 01). Targetscan Human7. 2 bioinformatics software predicted that the MTDH gene contained a target sequence of hsa-miR-26a.The luciferase reporter gene experiment also verified the targeted regulation relationship between miR-26a and MTDH. The expression of miR-26a was the highest in KYSE-450 cells and the lowest in Ecal09 cells. The mRNA expression of MTDH in the lv-miR-26a group was significantly lower than that in the lv-NC group, and the mRNA expression in the lv-miR-26a-inhibitor group was significantly higher than that in the lv-NC group. Alter tumor formation, miR-26a expression increased and MTDH expression decreased in miR-26a group. Alter tumor formation, the expression oi miR-26a decreased and the expression oi MTDH increased in miR-26a inhibitor group. Conclusion MiR-26a can inhibit the expression oi MTDH in esophageal cancer cells. Both in vitro and in vivo experiments can verily the targeted regulatory relationship between miR-26a and MTDH. MiR-26a ma)' play a role in the occurrence and development oi esophageal cancer through the MTDH.
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AIM: To optimize an orthopedic non-muscle invasive bladder cancer (NMIBC) model in nude mouse by comparing four different ways of cellular transplantation, and to evaluate the efficacy of drug by bladder instillation, so as to provide a stable and efficient animal model for the treatment of bladder cancer. METHODS: After disruption of bladder mucosa by dilute acid-alkali or silver nitrate, T24 cells were instilled into the nude mouse bladder. T24 cells were injected directly into the bladder with mechanical injury of bladder mucosa. T24 cells were injected into the bladder wall. On the 14th day after making models, the nude mice were sacrificed. And the bladder mass and histopathological changes of tumor (including bladder) was observe to confirm the formation of orthopedic bladder cancer. The dynamic changes of orthopedic bladder cancer were observed after injecting T24 cells into the bladder wall. Gemcitabine was used to verify the applicability of the model of injecting T24 cells into the bladder wall in vivo. RESULTS: No tumor was found in the bladder after intravesical instillation of T24 cells with dilute acid-alkali or silver nitrate treatment. With mechanical injury of bladder mucosa, all nude mice had tumors after injection T24 cells. But the number of tumors varied and often occurred at multiple sites. The tumor was found in the bladder of all nude mice by injecting T24 cells into bladder wall, and there was only one tumor. The tumor showed slow linear growth within 15 days and rapid linear growth from day 18 to 31. In vivo efficacy evaluation, gemcitabine 150 mg/kg intravesical perfusion could significantly inhibit the growth of NMIBC in nude mice replicated by direct injection of T24 cells into the bladder wall, and the tumor inhibition rate was 97.1%. CONCLUSION: The orthotopic NMIBC model can not be established with the bladder mucosa injuried by dilute acid-alkali or silver nitrate treatment. The number and size of orthotopic bladder cancer are different by mechanical injury of bladder mucosa. Injection of T24 cells into the bladder wall of nude mouse can successfully establish the orthotopic NMIBC model, which can be used for the evaluation of NMIBC therapeutic drugs.
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Objective:To investigate the effect of lncRNA UCA1 on the radiosensitivity of in vitro cultured glioma cell lines SHG-44, U87 and U251 by regulating the miR-873-5p expression. Methods:The survival of glioma cells SHG-44, U87 and U251 treated with different radiation intensities (0, 2, 4, 6 and 8 Gy) was detected by colony formation assay. The expression levels of UCA1 in glioma cells SHG-44, U87 and U251 were measured by qRT-PCR. The radiation-resistant glioma cells U87 and U251 were selected for subsequent study. After silencing UCA1 expression and/or over-expressing miR-873-5p, the cell survival rate was detected by colony formation assay, and the cell apoptosis rate was determined by flow cytometry. The dual luciferase reporter gene assay and qRT-PCR were employed to verify the targeting relationship between UCA1 and miR-873-5p.Results:UCA1 was up-regulated in the radiation-resistant U87 and U251 cells. Silencing UCA1 or over-expressing miR-873-5p inhibited the survival of U87 and U251 cells, and promoted the cell apoptosis induced by radiation exposure. miR-873-5p was a target gene of UCA1, and UCA1 negatively regulated the expression of miR-873-5p. The inhibition of miR-873-5p could reverse the effect of silencing UCA1 on the radiosensitivity of glioma cells. Silencing UCA1 increased the inhibitory effect of radiation on the glioma cell U251 xenografts.Conclusion:Silencing UCA1 inhibits the survival of glioma cells and promotes the cell apoptosis by up-regulating the expression of miR-873-5p, thereby increasing the radiosensitivity of glioma cells.
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【Objective】 To explore the establishment methods of transgenic human umbilical cord mesenchymal stem cells (hUC-MSCs) overexpressing tumor necrosis factor(TNF)-related apoptosis-inducing ligand (TRAIL) based on the transposons, and attempt to apply it on the nude mice mode with glioma. 【Methods】 PiggyBac transposon system specially designed by us was used to prepare non-targeting and Her2-targeting hUC-MSCs that can stably express TRAIL through puromycin screening. The glioma cells expressing firefly luciferase (U87MG-LUC) were injected into the skull of the immunodeficient mice (BALB/c-nu/nu) with 1×106 cells per mouse. After 7 days of injection, the mice transplanted with U87MG were detected with a small animal living imager to determine the size and location of the tumors in skull. Then we injected the glioma-transplantation nude mouse with two kinds of transgenic hUC-MSCs expressing TRAIL (named as untarget-TRAIL and target-TRAIL, respectively), or the non-transgenic hUC-MSCs (all 1×106 cells per mouse) or PBS (named as WT-MSCs and PBS for negative control) respectively, and then monitored the changes of tumor signals by a small animal living imager every week for 3~4 weeks. 【Results】 After six passages to expand the cells, the both transgenic cell lines can stably express TRAIL gene. Their ratio of green fluorescent protein (GFP) positive cells can reach 93%-97%, and the positive ratio of their MSC-specific surface markers still maintained normal (CD34+, CD45+, and HLA-DR+ all <0.1%, CD90>99%, CD73>88%, and CD105 >60%). The median survival time (d) of U87MG-transplanted nude mice in the groups of untarget-TRAIL, target-TRAIL, WT-MSCs, and PBS was 41 vs 39 vs 24 vs 23(P<0.05). 【Conclusion】 The transgenic hUC-MSCs overexpressing TRAIL gene can significantly prolong the survival time of nude mice with brain glioma.
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Objective To study the inhibitory effect of T lymphocytes secreting EphrinAl-Caspase-3 in vivo and on the growth of cancer cells in nude mice with breast cancer. Methods Nude mice (n = 35) were inoculated with breast cancer cells to construct a nude mouse model of breast cancer. When the tumor volume reached 0. 1 cm3, 30 nude mice with average size tumor tissue were randomly divided into PBS group, uninfected adenovirus group, T lymphocyte infected with Ad-EphrinAl-Caspase-3 group, and intratumoral transplantation. Tumor size was measured every day 2 to 3. Three groups of tumor-bearing nude mice were selected. After the above-mentioned cell transplantation, the subcutaneous tumor tissue homogenate was obtained every day 2 to 3, and the content of EphrinAl-Caspase-3 was detected by ELISA. At the end of the experiment, the animals in each group were sacrificed by cervical dissection and sliced. The presence of T lymphocytes expressing green fluorescent protein was observed under a fluorescence microscope, and Caspase-3 and Ki-67 were detected by immunofluorescence. Results After one week of inoculation of breast cancer cells into nude mice, the presence of subcutaneous tumors could be touched by hand, which proved that the tumor-bearing animals of breast cancer cells were successfully modeled. On the 8th day after inoculation, the tumor volume of the nude mice in each group became larger, and the difference between the treatment group and the PBS group/T lymphocyte group was extremely significant ( P<0.05). Although the tumor volume of the T lymphocyte transplantation group was slower than that of the PBS control group, there was no statistically significant difference between the two. The expression of EphrinAl-Caspase-3 was detected in the EphrinAl-Caspase-3 treatment group on the 2nd day, reached the peak on the 8th day, and then the secretion decreased gradually. No expression of EphrinAl-Caspase-3 was detected in the PBS control group and the T lymphocyte group. The presence of dispersed green fluorescent protein-labeled EphrinAl-Caspase-3-T lymphocytes was observed in the tumor tissues of the treatment group, while the presence of green fluorescent protein was not detected in the PBS group and the T lymphocyte groups. In the infected cells of the treatment group, the proportion of Caspase-3 positive cells was up- regulated, and the proportion of Ki-67 positive cells was down-regulated. No expression of EphrinAl-Caspase-3 was detected in the PBS group and the T lymphocyte group. Conclusion EphrinAl-Caspase-3 can significantly inhibit the growth of breast cancer cells, thereby exerting an anti-tumor effect.
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Objective: To investigate the effect of circulating estrogen level on the outcome of free fat grafting in nude mice. Methods: Eighteen female nude mice aged 6-8 weeks (weighing, 20-25 g) were randomly divided into 3 groups ( n=6). The nude mice in the ovariectomized group were treated with ovariectomy. The nude mice in the high estrogen group and the normal estrogen group only made the same incision to enter the peritoneum without ovariectomy. The nude mice in the high estrogen group were given the estradiol (0.2 mg/g) every 3 days for 30 days. The other two groups were given the same amount of PBS every 3 days. At 30 days after operation, the tail vein blood of nude mice in 3 groups were detected by estradiol ELISA kit, and the free fat (0.3 mL) donated by the females was injected into the sub-scalp of nude mice. After 8 weeks of fat grafting, the samples were taken for gross observation and weighing, and the prepared slices were stained with HE staining, CD31-perilipin fluorescence staining, immunohistochemical staining of uncoupling protein 1 (UCP1), and immunofluorescence staining of estrogen receptor α. The diameter of adipocytes and vascular density of adipose tissue were measured. The mRNA expressions of UCP1 and estrogen receptor α were detected by realtime fluorescence quantitative PCR (qRT-PCR). Results: All nude mice survived during experiment. ELISA test showed that the concentration of estradiol significantly decreased in the ovariectomized group and increased in the high estrogen group compared with the normal estrogen group ( P<0.05). At 8 weeks after fat grafting, the graft volume from large to small was ovariectomized group, normal estrogen group, and high estrogen group. There was significant difference in wet weight between the ovariectomized group and high estrogen group ( P<0.05). Section staining showed that compared with the normal estrogen group, the adipocytes in the ovariectomized group were larger, the expression of peri-lipoprotein was weaker, the vascular density decreased, and the expressions of UCP1 was negative, and the estrogen receptor α positive cells reduced. The above observation results in the high estrogen group were contrary to those in the ovariectomized group. There were significant differences in the diameter of adipocytes, the vascular density of adipose tissue, the number of the estrogen receptor α positive cells between groups ( P<0.05). The results of qRT-PCR showed that the mRNA expressions of UCP1 and estrogen receptor α significantly increased in the high estrogen group and decreased in the ovariectomized group compared with the normal estrogen group, and the differences were significant ( P<0.05). Conclusion: The level of circulating estrogen has a significant effect on the outcome of free fat grafting in nude mice. Low estrogen level leads to hypertrophy of graft adipocytes, while high estrogen level leads to the production of a large amount of beige fat and high vascular density in fat grafts, which may be related to the activation of estrogen receptor α on adipocytes.
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Objective To evaluate the effect of RNF2 gene knockdown in ECA109 cells on the radiosensitivity to esophageal cancer cell xenograft in nude mice. Methods Thirty-six male BALB/c/nu nude mice were randomly divided into 6 groups: control group, control+ irradiation group, NC group, NC+irradiation group, RNF2 shRNA group and RNF2 shRNA+ irradiation group. The nude mouse models with transplanted tumors were established by subcutaneous inoculation of EAC109 cells and given with irradiation at a dose of 3 Gy for 5 times. The longest ( a) and shortest ( b) diameters of the transplanted tumor were measured every 2 to 3 day since the fourteenth day after inoculation. The time of tumor formation was recorded. The tumor volume was calculated according to the formula ( ab2/2 ) . The growth curve was delineated. Three nude mice were sacrificed in each group at 24 h after the initial irradiation. The expression of RNF2 at the mRNA and protein levels in transplanted tumor tissues was measured by qRT-PCR and immunohistochemistry, respectively. The growth and tumor volume of the other nude mice in each group were observed. The cell apoptosis of transplanted tumor tissues was detected by TUNEL assay. The expression of Bcl-2 and Bax at the mRNA and protein levels in transplantated tumor tissues was quantitatively measured by qRT-PCR and immunohistochemistry, respectively. Results The tumor growth rate was the highest in the control and NC groups. The knockdown of RNF2 reduced the growth rate of xenografts and the tumor growth rate was the slowest in the RNF2 shRNA+ irradiation group ( P<0.05) . TUNEL assay revealed that the cell apoptosis rates in all groups were significantly increased after irradiation ( all P<0.05) . Before and after irradiation, the apoptosis rate in the RNF2 shRNA group was markedly higher than those in the control and NC groups ( both P<0.05) . Prior to irradiation, the expression levels of RNF2 mRNA and protein in the RNF2 shRNA group were significantly lower compared with those in the control and NC groups ( all P<0.05) , and the tendency became more significant after irradiation. Compared with the control and NC groups, the expression levels of Bcl-2 mRNA and protein were significantly down-regulated in the RNF2 shRNA group before and after irradiation ( all P<0.05) , whereas those of Bax mRNA and protein were considerably up-regulated ( all P<0.05 ) . Conclusions In vivo experiment demonstrates that RNF2 knockdown effectively increases the radiosensitivity of esophageal carcinoma EAC109 cells in nude mouse models with transplanted tumors, which is intimately associated with inducing the cell apoptosis.
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Objective@#To investigate the inhibitory effect of 17AAG-Cypate micelles on the non-small cell lung cancer A549 cells in nude mice and to explore its possible mechanism.@*Methods@#A549 lung adenocarcinoma tumor-bearing nude mice were established. The nude mice were treated with saline ( saline group), X-ray (X-ray group), 17AAG micelles+ X-ray (17AAG-M/X group) and 17AAG-Cypate micelles+ laser/X-ray (17AAG-Cypate-M/L+ X group), respectively. The growth of xenograft tumors in different groups was measured on a regular basis to delineate the growth curve. The expression of proliferating cell nuclear antigen (PCNA) was measured by immunohistochemistry. The microvascular density was detected. The apoptosis of xenograft tissues was observed by TUNEL staining. The expression levels of p-ERK1/2 and p-AKT were quantitatively measured by Western blot.@*Results@#Compared with the saline group, varying degrees of inhibition of tumor growth were observed in the X-ray, 17AAG-M/X-ray and 17AAG-Cypate-M/L+ X groups, particularly in the 17AAG-Cypate-M/L+ X group (all P<0.05). In all groups, the expression levels of PCNA were significantly down-regulated (all P<0.05), the microvascular density was remarkably reduced (all P<0.05) and the expression levels of p-ERK1/2 and p-AKT were considerably down-regulated (all P<0.05).@*Conclusions@#17AAG-Cypate micelles can inhibit the growth of human non-small cell lung cancer in nude mice, probably by reducing the activity of p-ERK1/2 and p-AKT, thereby weakening the activation of the MAPK-ERK and PI3K-AKT signaling pathways.
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Objective: To observe the growth inhibition effect of Aconiti Lateralis Radix Praeparata polysaccharide on the gastric cancer xenografts in nude mice and the expressions of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-14 (MMP-14) in tumor tissues. Method: The nude mice xenograft model was established and randomly divided into model group, 5-fluorouracil group (5-FU, 0.01 g · kg-1), high and low-dose Aconiti Lateralis Radix Praeparata polysaccharide groups (0.2, 0.1 g · kg-1). Each group was given drug by gavage for 15 days. The effect of Aconiti Lateralis Radix Praeparata polysaccharide on the weight of gastric cancer in nude mice was observed. Morphological changes of tumor cells were observed under light microscope. The content of transforming growth factor-β1 (TGF-β1) in serum was determined by enzyme-linked immunosorbent assay (ELISA). The protein and mRNA expressions of MMP-2, MMP-14 in tumor tissues were detected by immunohistochemistry, Western blot and Real-time PCR. Result: The tumor inhibition rates of high and low-dose Aconiti Lateralis Radix Praeparata polysaccharides were 52.83%,40.57%, respectively. Compared with model group, the tumor weight of high and low-dose Aconiti Lateralis Radix Praeparata polysaccharide was significantly lower (Pβ1 content in nude mice (PPPPPPPConclusion: Aconiti Lateralis Radix Praeparata polysaccharide can inhibit the growth of gastric cancer xenografts in nude mice and the expression of TGF-β1.The anti-tumor mechanism may be related to the down-regulation of MMP-2, MMP-14 protein and mRNA expressions.
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Objective@#To investigate the effect and mechanism of LncRNA ANRIL on the radiosensitivity of HCT116 cells line and nude mouse transplant tumors.@*Methods@#The expression of LncRNA ANRIL in colorectal cancer cells was detected by qPCR. The negative control siRNA, ANRIL siRNA, miR-NC mimic, miR-195 mimic, miR-NC inhibitor and miR-195 inhibitor were transfected into HCT116 cells, and marked as negative control group, silencing ANRIL group, overexpressing miR-NC group, overexpressing miR-195 group, inhibiting miR-NC group and inhibiting miR-195 group, and the HCT116 cells without any treatment were marked as the blank control group. The clone formation assay was used to detect radiosensitivity of colorectal cancer cells, flow cytometry was used to detect apoptosis. The web site, StarBase, was used to predict the downstream miRNAs of ANRIL and dual luciferase reporter gene assay was used to further verify. Subcutaneous tumor transplantation assay was used to detect the effect of ANRIL on the growth of colorectal cancer cells after irradiation.@*Results@#After irradiation with 2, 4, 6 and 8 Gy, the cell survival fraction of silencing ANRIL group was significantly decreased when compared with that of negative control group (P<0.05), and the radiosensitivity ratio was 1.52. The apoptosis rate of the silencing ANRIL+ 4 Gy group was significantly higher than that of the negative control+ 4 Gy group ((27.86±2.78)% vs. (12.06±1.46)%, P<0.05). The results of the experiment on nude mouse transplant tumors showed that the tumor volume in the negative control group was lower than that of the silent ANRIL group on days 13, 16, 19, 22 and 25 ((234±66) mm3, (273±63) mm3, (296±72) mm3, (321±85) mm3 and (403±94) mm3 vs. (357±79) mm3, (485±124) mm3, (617±143) mm3, (764±174) mm3 and (985±221) mm3P<0.05). MiR-195 is a target gene of ANRIL, and inhibition of miR-195 can reverse the inhibitory effect of silencing ANRIL on radiosensitivity, apoptosis and xenografts of HCT116 cells.@*Conclusions@#LncRNA ANRIL regulates the radiosensitivity of colorectal cancer cells by miR-195, which may provide a new sensitizing target for clinical colorectal cancer radiotherapy.
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We study here in vitro cytotoxicity, in vivo tumor inhibition and the mechanism on photodynamic therapy (PDT) of photosensitizer BF01 using human hepatocellular carcinoma cell line BEL-7402. CCK-8 method was used to detect the inhibition rate and IC50 in BEL-7402 cells on the same laser intensity with varying concentrations (0, 0.8, 1.6, 3.2, 6.4 μmol·L-1) of photosensitizer BF01. Cell death mode of BEL-7402 was detected by flow cytometry, with apoptotic characteristics observed by DAPI staining, and the subcellular localization of reactive oxygen was observed using photodynamic detection and confocal microscopy. The cell model of human liver cancer in nude mice was established, tumor growth curve was drawn, and the therapeutic effect of BF01 was determined. The animal experimentation was approved by East China University of Science and Technology Ethics Committee. The results indicated that BF01 PDT treatment can clearly inhibit BEL-7402 tumor cell proliferation, with the killing rate of 86% at the concentration of 6.4 μmol·L-1 of BF01, and half lethal concentration IC50 value of 2.46 μmol·L-1. DAPI stained nuclei shows the characteristics of advanced stage apoptosis, whereas reactive oxygen species level in the mitochondria increased with increasing drug concentration. In vivo experiments showed that photosensitizer BF01 mediated photodynamic therapy of liver cancer cells and inhibited tumor growth in mice. Therefore, the new BF01 photosensitizer has a potential for development into future clinic application.
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Objective: To investigate the effects of adipose-derived stem cells (ADSCs) and endothelial cells (ECs) on the survival and neovascularization of fat tissue transplants. Methods: The ADSCs were isolated by collagenase digestion from the adipose tissues voluntarily donated by the patients undergoing mastectomy, and subcultured. The passage 3 ADSCs were used for subsequent experiments. The residual fat tissues were used to prepare fat particles (FPs). The human umbilical vein endothelial cells (HUVECs) were used as ECs for subsequent experiments. Eighty healthy male nude mice, aged 4-6 weeks, were randomly divided into 4 groups ( n=20). The mice were received subcutaneous injection at the dorsum of 1 mL FPs+0.3 mL normal saline (NS) in control group, 1 mL FPs+2×10 6 ECs+0.3 mL NS in ECs group, 1 mL FPs+2×10 6 ADSCs+0.3 mL NS in ADSCs group, and 1 mL FPs+1×10 6 ECs+1×10 6 ADSCs+0.3 NS in ADSCs+ECs group. General observations of the injection sites were performed, and the survival of the mice was recorded. At 2, 4, 8, and 12 weeks after injection, grafted fat tissues were firstly assessed by ultrasonography, then they were collected for volume measurement (water displacement method) and histology observation (HE staining and immunofluorescence staining). Results: All mice survived until the end of experiment. At each time point, no significant difference was noted between groups in ultrasonography assay. There was no significant blood flow signal in the grafted fat tissues, or cysts, calcification, solid occupying in recipient area. Generally, the volume of grafted fat tissues decreased with time in all groups. Specifically, the volumes of grafted fat tissues were larger in ADSCs group and ADSCs+ECs group than that in control group and ECs group ( P<0.05) at each time point, and in ADSCs group than in ADSCs+ECs group ( P<0.05) at 8 and 12 weeks. HE staining showed that all groups had similar tendencies in general histology changes, and remodeling in ADSCs group was the fastest than in the other groups. By immunofluorescence staining for neovascularization, the new vessels in all groups were increasing with time. The vessel densities were higher in ECs group, ADSCs group, and ADSCs+ECs group than in control group ( P<0.05) at each time point, in ADSCs group than in ECs group and ADSCs+ECs group ( P<0.05) at 4 weeks, in ADSCs group and ADSCs+ECs group than in ECs group ( P<0.05) at 8 and 12 weeks. Conclusion: ADSCs can significantly increase the survival of transplanted fat tissue, which may be related to promoting the neovascularization.
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Objective To investigate the effects of knockdown of miR-449a on breast cancer xenograft in nude mice.Methods In this study,we established a model of breast cancer xenograft in nude mice with breast cancer MCF-7 cell line,then stably transfected MCF-7 cells with plasmids containing shRNA-miR-449a precursors;the stable transfected cells were injected subcutaneously into the nude mice by multi-point injection.We used negative plasmid as negative control and PBS as blank control.We detected tumor volume after the treatment of nude mice and tumor growth inhibition rate was calculated.The expressions of miR-449a and Notch1 in xenograft in nude mice were detected by real-time PCR;the levels of Notch 1,β-catenin and E-cadherin in the xenograft tumor tissues were detected by Western blot; and the expression of E-cadherin in tumor tissues was detected by immunohistochemistry.Results In the process of xenograft tumor formation,no nude mice died.Knockdown of miR-449a could significantly inhibit the growth of tumor volume after tumor detection (P <0.05 ).miR-449a was significantly down-regulated in xenograft tumor tissues,and the expressions of Notch 1 and β-catenin were also down-regulated,while the level of E-cadherin was increased.The results of immunohistochemistry showed that the expression of Notch1 protein was decreased and that of E-cadherin protein was increased (P <0.05).Conclusion Knockdown of miR-449a inhibited the normal growth of breast cancer xenografts in nude mice by down-regulating the expressions of Notch 1 andβ-catenin and promoting the expression of E-cadherin.
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Objective The purpose of this study was to compare the adenomyosis models in nude mice generated by four different methods,and to find out an optimal modeling method, and to provide an ideal animal model for exploring pathogenesis and experimental treatment of uterine adenomyosis. Methods 1. 80 female healthy nude mouse were divided randomly into 4 groups: Intraperitoneal implantation group, subcutaneous implantation group, intraperitoneal injection group, and subcutaneous injection group. The transplants were taken for pathological examination at 4 weeks after surgery. Results The success rate of intraperitoneal implantation group was 95%,and that of the subcutaneous implantation group was 45%,while the success rate of intraperitoneal injection group and subcutaneous injection group was 0%. Conclusions Establishment of a nude mouse model of uterine adenomyosis by intraperitoneal implantation method has a high success rate and with good stability, and is an ideal mouse model of human-derived uterine adenomyosis.
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Objective · To prepare the liposomal pemetrexed and investigate its effects on MCF-7 breast cancer cells in vitro and nude mice bearing MCF-7 xenograft tumors. Methods · Liposomal pemetrexed was prepared by film dispersion method. Inhibition of MCF-7 breast cancer cell lines was evaluated by CCK-8 method, and anti-tumor effects were investigated on Balb/c nude mice bearing MCF-7 xenograft tumors. Results · Liposomal pemetrexed inhibited the growth of MCF-7 cells. When the concentrations of pemetrexed were 0.20, 0.40 and 10.00 μg/mL, the cell viability in experiment group (liposomal pemetrexed) was significantly lower than that in control group (pemetrexed of same concentration gradient), with P values of 0.013, 0.035 and 0.041, respectively. Compared with blank group (same volume of PBS), the volumes and weights of tumors of nude mice in experiment group(liposomal pemetrexed) and control group (same volume of pemetrexed) were significantly lower, and the volume and weight of tumor in experiment group were also significantly lower than those in control group (P=0.000). Conclusion · Compared to bulk drug of pemetrexed, liposomal pemetrexed can inhibit the growth of MCF-7 breast cancer cells and the Balb/c nude mice bearing MCF-7 xenograft tumors.
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Objective To investigate the effects of knockdown of miR-449a on breast cancer xenograft in nude mice.Methods In this study,we established a model of breast cancer xenograft in nude mice with breast cancer MCF-7 cell line,then stably transfected MCF-7 cells with plasmids containing shRNA-miR-449a precursors;the stable transfected cells were injected subcutaneously into the nude mice by multi-point injection.We used negative plasmid as negative control and PBS as blank control.We detected tumor volume after the treatment of nude mice and tumor growth inhibition rate was calculated.The expressions of miR-449a and Notch1 in xenograft in nude mice were detected by real-time PCR;the levels of Notch 1,β-catenin and E-cadherin in the xenograft tumor tissues were detected by Western blot; and the expression of E-cadherin in tumor tissues was detected by immunohistochemistry.Results In the process of xenograft tumor formation,no nude mice died.Knockdown of miR-449a could significantly inhibit the growth of tumor volume after tumor detection (P <0.05 ).miR-449a was significantly down-regulated in xenograft tumor tissues,and the expressions of Notch 1 and β-catenin were also down-regulated,while the level of E-cadherin was increased.The results of immunohistochemistry showed that the expression of Notch1 protein was decreased and that of E-cadherin protein was increased (P <0.05).Conclusion Knockdown of miR-449a inhibited the normal growth of breast cancer xenografts in nude mice by down-regulating the expressions of Notch 1 andβ-catenin and promoting the expression of E-cadherin.
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Objective To observe the effects of TLR9 on the nude mouse transplanted tumor growth of human pancreanc cancer and its drug resistance.Methods The nude mouse transplated tumor of human pancreatic cancer PANC-1 was established and randomly divided into 6 groups for conducting the experiment:sterile normal saline group,TLR9 agonist,TLR9 inhibitor group,gemcitabine group,TLR9 inhibitor plus gemcitabine Bin group,TLR9 agonist plus gemcitabine.The tumor size and growth situation were recorded by the vernier caliper.The immunohistochemical method was used to detect tumor TLR9 receptor expression.The tumor growth,metastasis and paracancerous nssue invasion situation were observed by the magnetic resonance imaging (MRI).Results The volume and growth speed of resected tumor mass in the gemcitabine group,TLR9 agonist + gemcitabine group,TLR9 inhibitor plus gemcitabine group was significantly smaller than those in other groups (P＜0.05),which in the TLR9 agonist + gemcitabine group were significantly greater than those in the TLR9 inhibitor plus gemcitabine group and gemcitabine group (P＜0.05),the difference between the TLR9 inhibitor plus gemcitabine group and gemcitabine group had statistical significance (P＜0.05),while the difference among the TLR9 agonist group,TLR9 inhibitor group and normal saline group had no stastistical significance (P＞0.05).The tumor in mice at 7 weeks after planting showed oval shape with clear boundary by MRI observation,no obvious metastais and paracancerous invasion were seen in paracancerous nssues no statistically significant,5 weeks,6 weeks after planting,seven weeks mice observed in MRI,the tumor into an,state clearly that the transfer of the surrounding tissue,no significant vascular invasion,heart,liver,kidney disease.The TLR9 expression on the surface of tumor tissue was detected and identified.Conclusion Pancreatic cancer nude mouse transplated tumor has definitely positive expression of TLR9,TLR9 activation can significantly decrease the sensitivity of pancreatic cancer to chemotherapy,increases the drug resistance of tumor,on contrary promotes the tumor growth.
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Objective To explore the effect of adriamycin on the characteristics of colony derived from human adrenal cortical carcinoma cells (ACC) SW-13.Methods Treatment with Adriamycin (ADM) was used in BALB/c-nude mouse tumor xenograft model established using the ACC cell line SW-13.The characteristic of colony was assessed for the formation rates,the percentagc of three colony types and growth curve of single cell.Hoechst33342 dyeing test was used to test drug resistance.Results The Single-cell colony formation rate of experimental group were significantly higher than control group (P ＜ 0.05),and the holoclone percentage of experimental group were significantly higher than control group (P ＜ 0.05).In the Hoechst33342 dyeing tcst,the fluorescence intensity of control was higher than experimental group.Conclusion The treatment of ADM in vivo is beneficial for the colony formations of ACC cell and the formations rate of holoclone,and can improve the ability of drug resistance of ACC cell SW-13.