Quantitative analysis of proteins related to chemoresistance to paclitaxel and carboplatin in human SiHa cervical cancer cells via iTRAQ / 부인종양

Objective@#This study aimed to identify proteins related to paclitaxel and carboplatin chemoresistance in cervical cancer. @*Methods@#Quantitative proteomic analysis was performed on normal SiHa cells and those treated with paclitaxel and carboplatin for 14 days, with isobaric tags for relative and absolute quantitation (iTRAQ) analysis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were used to identify related processes and differentially expressed proteins. @*Results@#A total of 67 and 96 differentially expressed proteins were identified in the paclitaxel- and carboplatin- treated groups, respectively. GO and KEGG enrichment analyses identified 53 (43 upregulated and 10 downregulated) and 85 differentially expressed proteins (70 upregulated and 15 downregulated) in the paclitaxel- and carboplatin-treated groups, respectively. The cell counting kit-8 results revealed that APOA1 was overexpressed in both the paclitaxel- and carboplatin- resistant SiHa cells compared with the control cells. Immunohistochemistry showed that APOA1 was highly expressed in the paclitaxel- and carboplatin- resistant squamous cell carcinoma of the cervix. @*Conclusion@#This study is the first to use iTRAQ to identify paclitaxel- and carboplatin- resistance proteins in cervical cells. We identified several proteins previously unassociated with paclitaxel and carboplatin resistance in cervical cancer, thereby expanding our understanding of paclitaxel and carboplatin resistance mechanisms. Moreover, these findings indicate that the APOA1 protein could serve as a potential marker for monitoring and predicting paclitaxel and carboplatin resistance levels.

Effect of tamoxifen on endometrium of patients after breast cancer operation / 中国实用妇科与产科杂志

OBJECTIVE: To investigate the clinicopathological effects of taking tamoxifen(TAM)on endometrium after breast cancer operation.METHODS: From January 2011 to December 2017,622 cases of breast cancer were treated in Beijing Obstetrics and Gynecology Hospital,Capital Medical University.Among them,197 patients took TAM,and 59 patients who took TAM underwent hysteroscopic surgery due to abnormal vaginal bleeding or ultrasound endometrial abnormalities.The 59 patients were divided into premenopausal and postmenopausal groups to analyze the pathological condition;the medication time was defined as 3 years and 5 years,so as to observe the endometrial pathology.Set the endometrial abnormal hyperplasia includes: endometrial cancer and the endometrial atypical hyperplasia, and the rest are benign endometrial lesions and the normal endometrium, and then analyze the ultrasound criteria for abnormal endometrial thickening in premenopausal and postmenopausal according to the endometrial pathology.RESULTS: Among the 197 patients who took TAM after breast cancer,59 patients underwent hysteroscopic surgery,32.2%(19/59)of them visited the hospital because of abnormal vaginal bleeding,76.3%(45/59)were pathologically confirmed to have a lesion,in which the endometrial polyps was with the highest incidence.The incidence of endometrial cancer after menopause was 20.0%(6/30),premenopausal endometrial cancer 3.4%(1/29),and atypical hyperplasia before menopause was 20.7%(6/29).When taking TAM for more than 3 or 5 years,the incidence of endometrial cancer and atypical hyperplasia increased.Premenopausal ultrasound endometrial thickness is associated with abnormal endometrial hyperplasia(P=0.035).The endometrial thickness 15 mm can be used as the best diagnostic ultrasound cut-off for the diagnosis of premenopausal abnormal endometrial thickening. Postmenopausal ultrasound endometrial thickness was not associated with abnormal endometrial hyperplasia(P=0.631).CONCLUSION: Taking TAM after breast cancer surgery can result in endometrial polyps and endometrial hyperplasia.Premenopausal patients can also have endometrial cancer and atypical hyperplasia,so the endometrial monitoring should not be ignored.Those who take TAM for more than 3 years need to be more alert to the occurrence of endometrial lesions.The premenopausal B-ultrasound endometrial thickness 15 mm can be used as the best diagnostic ultrasound cut-off for the diagnosis of abnormal endometrial thickening. After the menopause, the endometrial thickness of 5 mm was still used as the standard for abnormal endometrial thickening.

Research on monoclonal antibody OX26 modified salvianolic acid B/baicalin nanostructured lipid carriers / 中草药

Objective To prepare, characterize, and study cellular uptake of transferrin receptor monoclonal antibody OX26 modified nanostructured lipid carrier loaded with salvianolic acid B and baicalin (Sal B/BA-NLC). Methods Sal B/BA-NLC was prepared by emulsification-solvent evaporation method. OX26 was thiolated with 2-iminothiolane hydrochloride and then conjugated to the surface of Sal B/BA-NLC. The morphology, particle size, Zeta potential, and encapsulation efficiency (EE) were evaluated for the physicochemical properties, and OX26 modified Sal B/BA-NLC was verified by differential scanning calorimetry (DSC) and nuclear magnetic resonance spectroscopy (NMR). Coumarin-6 (C6) was used as the fluorescent probe instead of baicalin and salvianolic acid B to prepare the formulations in cellular uptake study. The cellular uptake study was conducted by brain microvascular endothelial cells bEnd.3 using high content cell imaging analysis system. Results The prepared OX26 modified Sal B/BA-NLC had particle size of (27.50 ± 3.37) nm, PDI of 0.39 ± 0.04, and Zeta potential of (-7.06 ± 1.85) mV. The DSC and NMR results indicated that the drug was encapsulated in the nanostructured lipid carrier in an amorphous form. The results of cell uptake showed that the fluorescence intensities of the three solutions in bEnd.3 cells were: OX26-C6-NLC > C6-NLC > C6-SL. Conclusion The prepared OX26 modified Sal B/BA-NLC has smaller particle size, uniform distribution, and high EE. The OX26-modified NLC group had a higher intake than the solution group and the unmodified NLC group.

Pharmacokinetics of bullatine A in Aconitum brachypodum total alkaloids gel in transdermal delivery / 中国中药杂志

In this study, the changes of bullatine A in plasma and skin of mice with time in microemulsion gel and ordinary gel of Aconitum brachypodum total alkaloids were compared through UPLC-MS/MS, and their pharmacokinetic parameters were also compared and analyzed, to investigate the feasibility of microemulsion agent in the transdermal drug delivery. UPLC-MS/MS method for simultaneous determination of bullatine A in plasma and skin had high sensitivity and was in line with the pharmacokinetic study requirements for transdermal drug delivery. The main pharmacokinetic parameters for microemulsion gel in the plasma were as follows: Cmax=(37.62±14.31) μg•L⁻¹, Tmax=(3.40±1.34) h, AUC0-∞=(1 027.7±260) μg•L⁻¹•h⁻¹, MRT=(34.80±12.31) h, MRTlast=(10.68±0.57) h, t1/2=(23.11±9.20) h; main pharmacokinetic parameters for ordinary gel in the blood: Cmax=(52.23±15.90) μg•L⁻¹, Tmax=(4.00±0.00) h, AUC0-∞=(728.60±280.80) μg•L⁻¹•h⁻¹, MRT=(20.69±3.98) h, MRTlast=(9.34±0.42) h, t1/2=(14.69±3.15) h. The results showed that the microemulsion gel had more stable transdermal absorption, longer duration of action and higher bioavailability than ordinary gel, indicating that the microemulsion gel had a good and stable transdermal effect. There was no significant difference in bioavailability of bullatine A in skin between microemulsion gel and ordinary gel.

In vitro recovery rate of bullatine A microdialysis probe / 中国中药杂志

To establish UPLC-MS/MS method for determination of the recovery rate of bullatine A microdialysis probe. The concentration difference method(incremental method, decrement method) was used to measure in vitro recoveries, and the effects of perfusate pH value, flow rate, concentration, and temperature on the recovery rate were investigated to explore the feasibility of microdialysis for the pharmacokinetic study of bullatine A. The method of UPLC-MS/MS showed good linear relationship within the required range; the specificity, recovery rate and precision of chromatography met the requirements of microdialysis samples. There was no significant difference in the measured recovery rate between incremental method and decrement method. Under the same conditions, in vitro recovery rate of the probe was decreased with the increase of flow rate, and was significantly increased with the increase of temperature, but was independent of bullatine A concentrations around the probe. The results showed that, microdialysis technology can be used for the pharmacokinetic study of bullatine A, and retrodialysis method (decrement method) can be used for the determination of the in vivo recovery rate of bullatine A microdialysis.

A Population-based Study of Invasive Cervical Cancer Patients in Beijing: 1993-2008 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>In recent years, the incidence of cervical cancer has been rising, particularly in young adults, as the second most common gynecological cancer in China. The aim of this study was to explore the incidence change and the epidemiological characteristics of cervical cancer in Beijing over the past 16 years.</p><p><b>METHODS</b>All the cases of the study were limited to Beijing residents diagnosed with cervical cancer and registered in Beijing from January 1, 1993, to December 31, 2008. A total of 4100 patients with cervical cancer were obtained from the Statistics Database of Beijing Cancer Registry (BJCaR). According to the registered data, we retrospectively reviewed all original cases which we can acquired in reported hospital. Cervical situ cancer, cervical metastatic cancer, non-Beijing residents and repeatedly registered cases were excluded. Totally, 3641 registered cases were verified correctly. Meanwhile, we also collected the following data: Age, occupation, detected methods, histological type, and staging. The trends of incidence and mortality were analyzed by Joinpoint Regression Program 4.1.1.1 produced by National Cancer Institute (NCI, USA). The annual percent change (APC) was calculated using the Joinpoint regression model.</p><p><b>RESULTS</b>The crude rates of incidence and mortality were 10.4 and 1.0 per 100,000 women, respectively during 1993 to 2008. The average WHO age-standardized incidence rates were 11.5 per 100,000 women. There was a decrease in incidence annually by 8.0% (P = 0.3) during 1993-1996 and a rapid increase annually by 18.9% after 1999 (P < 0.01). The median age was 67 years in 1993, but the median age decreased to 45 years in 2008. The peak of the age-specific incidence curve was at 40 years in the most recent period (2005-2008), which was 25-30 years earlier than that in previous periods (1993-1996). In the 2224 cases, the numbers of patients with stage I, II, III and IV were 910 (40.9%), 601 (27%), 542 (24.4%), 171 (7.7%), respectively. The percentage of patients with stage I was 7.6% (13/171) in 1993-1996, but the percentage increased to 51.6% (643/1247) in 2005-2008 (P < 0.01). Otherwise the percentage of advanced stage (stage III-IV) during the same period was dropped down significantly from 52.0% (89/171) to 22.5% (280/1247) (P < 0.01). Unemployed and housewife ranked first accounting for 27.3% of the total (607/2224). Urban low-income people such as worker ranked the second accounting for 17.0% (377/2224), the third place was farmer accounting for 14.0% (312/2224). Only 381 (17.1%, 381/2224) women in 2224 were first detected cervical cancer by routine screenings. Company staff (36.5%, 139/381), professional and technical personnel (22.6%, 86/381), national official (22.0%, 84/381) occupied the top three (total 81.1%) in the 381 patients detected cervical cancer by screening.</p><p><b>CONCLUSIONS</b>The cervical cancer incidence has showed a continuous rise in Beijing since 1999. Government-led mass screening should target the low socioeconomic population primarily. Meanwhile the government should enhance public health education of cancer screening to increase the rate of screening.</p>

Analysis of differentially expressed proteins in normal cervix, cervical intraepithelial neoplasia and cervical squamous carcinoma tissues / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To explore the differentially expressed proteins in normal cervix, cervical intraepithelial neoplasia (CIN) and squamous cervical carcinoma (SCC) tissues by differential proteomics, and to provide a basis for studies on CIN molecular pathogenesis, clinical diagnosis and treatment.</p><p><b>METHODS</b>Uterine cervical tissue specimens from the patients treated between August 2008 and September 2009 in the Department of Oncology of Beijing Obstetrics and Gynecology Hospital were collected. There were samples of normal cervix (n = 9), CIN (n = 23, CIN I = 7, CIN II = 8, CIN III = 8) and SCC (n = 7). 2-D DIGE and DeCyder software were used to detect the differentially expressed protein-spots. Then MALDI-TOF/TOF MS was used to analyze the differentially expressed proteins. Collect normal cervix(n = 20), CIN (n = 60) and SCC (n = 20), immunohistochemistry (IHC) and Western blot were used to verify the differentially expressed proteins of S100A9 (S100 calcium-binding protein A9) , eEF1A1 (eukaryotic elongation factor 1-alpha-1) and PKM2 (pyruvate kinase isozymes M2) among the normal cervix, CIN and SCC tissues. Immunohistochemistry was used to detect the differentially expressed S100A9, eEF1A1 and PKM2 in the cervical tissues.</p><p><b>RESULTS</b>2D gel electrophoresis images with high resolution and good repeatability were obtained. Forty-six differentially expressed proteins (27 were up-regulated and 19 were down-regulated) were selected among the normal, CIN, and SCC, and 26 proteins were successfully identified. Immunohistochemistry showed that protein S100A9 was mainly expressed in the cytoplasm, and its positive expression rate was 20.0% in normal cervical mucosa, 70.0% in CIN, and 100.0% in squamous cell carcinoma, with a significant difference between them (P = 0.006). eEF1A1 was mainly expressed in the cell plasma. Its positive expression rate was 70.0% in normal cervix, 73.3%in CIN and 60.0% in SCC tissues, with a non-significant difference between them (P = 0.758). The protein PKM2 was mainly expressed in the cell nuclei. Its positive expression rate was 100.0% in normal cervix, 93.3% in CIN and 75.0% in SCC tissues, showing a difference close to statistical significance (P = 0.059) between them. The results of Western blot were similar with that of immunohistochemical examination.</p><p><b>CONCLUSIONS</b>There are differentially expressed proteins among normal cervix, CIN and SCC. S100A9, eEF1A1 and PKM2 may become candidate markers for early diagnosis of cervical cancer and new targets for therapy. It also provides a further basis for studies of the pathogenetic mechanism of CIN developing to cervical cancer.</p>

Association of MMP14 gene polymorphisms and osteoporosis in Zhuang men from Baise region of Guangxi / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To assess the association between membrane type 1 matrix metalloproteinase gene (MT1-MMP, MMP14) polymorphisms and osteoporosis in Zhuang men from Baise region of Guangxi.</p><p><b>METHODS</b>Genotypes of 5 loci (rs1003349, rs3751488, rs2269213, rs2236303 and rs743257) of MMP14 gene in 301 Zhuang men were determined with single base extension methods, and bone mineral density (BMD) at left calcaneus was evaluated with quantitative ultrasound with measured values of broadband ultrasonic attenuation (BUA). The subjects were divided according to BMD into osteoporosis group, osteopenia group and normal bone density group.</p><p><b>RESULTS</b>All selected loci were in Hardy-Weinberg equilibrium (P> 0.05). By multiple linear stepwise regression analysis, polymorphisms of the five loci were not associated with BUA. But a significant higher risk of osteoporosis was found in individuals with MMP14 rs1003349 GT genotype (vs. GG genotype; P<0.05) and rs2236303 CC and CT genotypes (vs. TT genotype; P<0.05). Genetic linkage between rs1003349 and rs2236303 was also discovered (D'= 0.839, r(2) = 0.458, P<0.01). Compared with the normal bone density group, the frequency of a G-T haplotype of rs1003349 and rs2236303 was significantly lower in the osteoporosis group (P<0.05). And the risk of osteoporosis for individuals with G-C and T-C haplotypes was 2.556 (95% CI: 1.029-6.349, P = 0.038) and 5.111 (95% CI: 1.341-19.485, P = 0.011) compared with G-T haplotype.</p><p><b>CONCLUSION</b>Polymorphisms of rs1003349 and rs2236303 loci of MMP14 gene are associated with the susceptibility of osteoporosis in Zhuang men in Guangxi. G-C and T-C haplotypes for loci rs1003349 and rs2236303 may increase the disease risk.</p>

Gain of human telomerase RNA gene is associated with progression of cervical intraepithelial neoplasia grade I or II / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The 3q26 chromosome region, where the human telomerase RNA gene (hTERC) is located, is a biomarker for cervical cancer and precancerous lesions. The aim of this study was to confirm the value of measuring hTERC gene gain in predicting the progression of cervical intraepithelial neoplasia grade I or II (CIN-I and -II, respectively) to CIN-III and cervical cancer.</p><p><b>METHODS</b>Liquid-based cytological samples from 54 patients with CIN-I or CIN-II lesions were enrolled in this study. Follow-up was performed with colposcopy and biopsy within 24 months after the diagnosis of CIN-I or CIN-II. Copy numbers of the hTERC gene were measured by fluorescence in situ hybridization with a dual-color probe mix containing the hTERC gene probe (labeled red) and the control, the chromosome 3 centromere-specific probe (labeled green).</p><p><b>RESULTS</b>All patients whose lesions progressed from CIN-I or CIN-II to CIN-III displayed a gain of the hTERC gene, whereas patients where the hTERC gene was not amplified did not subsequently progress to CIN-III or cervical cancer. The signal ratio pattern per cell was recorded as N:N (green:red). The numbers of cells with the signal ratio pattern of 4:4 or N:≥5 in patients whose lesions progressed to CIN-III were significantly higher than those whose lesions did not progress. Significantly, none of the patients with a 4:4 signal ratio pattern regressed spontaneously.</p><p><b>CONCLUSIONS</b>In conclusion, measurement of hTERC gene gain in CIN-I or CIN-II patients using liquid-based cytological samples could be a useful biomarker to predict the progression of such cervical lesions. In addition, a 4:4 or N:≥5 signal ratio pattern may indicate the unlikeness of spontaneous regression of CIN-I or CIN-II lesions.</p>

Analysis of serum biomarkers of ovarian epithelial cancers based on 2-DE DIGE and MALDI TOF/TOF / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To find new serum tumor markers for ovarian epithelial cancers by 2-DE DIGE and MALDI-TOF/TOF proteomic methods, in order to improve the diagnostic sensitivity and specificity.</p><p><b>METHODS</b>Serum samples from 103 cases of ovarian epithelial cancers, 60 cases of healthy women, 63 cases of benign ovarian tumors and 63 cases of benign pelvic diseases were collected. Sera of 20 cases of ovarian epithelial cancers (A), 20 cases of ovarian benign tumors (B), 20 cases of pelvic benign diseases (C) and 20 cases of health control (D) were matched by age and pooled, respectively. After depletion of high abundance serum albumin and IgG, the samples were assayed by 2-DE DIGE. The test was repeated three times. Analysis with DeCyder software revealed significant differential protein spots which were identified by MAIDI-TOF/TOF. Western blot and ELISA were used to validate the candidate serum markers.</p><p><b>RESULTS</b>1) There were 41 proteins having significant differences between the groups. MAIDI-TOF/TOF successfully identified 28 proteins. Haptoglobin (Hp) was the most significantly up-regulated protein, and transferrin (Tf) was the most significantly down-regulated protein. 2) Western blot and ELISA proved that there were significant differences in Hp and Tf between ovarian epithelial cancers and normal controls (P = 0.000), between ovarian epithelial cancers and ovarian benign tumors (P = 0.000), between ovarian epithelial cancers and benign pelvic disease sera (P = 0.000). 3) CA125 + Hp + Tf combined detection of ovarian cancer had higher sensitivity and specificity than CA125, Hp or Tf detection alone.</p><p><b>CONCLUSION</b>Hp and Tf are differently expressed in the sera of patients with ovarian epitheliual cancers. They can be used as serum biomarkers for ovarian epithelial cancers. CA125 + Hp + Tf combined detection may improve the sensitivity and specificity of diagnosis of ovarian epithelial cancers.</p>

Detection of differentially expressed genes in human autosomal dominant polycystic kidney tissue / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To detect the differentially expressed genes in human polycystic kidney by cDNA microarray.</p><p><b>METHODS</b>The PCR products of 8398 genes were spotted onto a chip in array. Both mRNAs isolated from polycystic kidney tissue and normal kidney tissue were reversely transcribed to cDNAs with the incorporation of fluorescent dUTP (Cy5-dUTP and Cy3-dUTP) for preparing the hybridization probes. The mixed probes were hybridized to the cDNA microarray. Then the cDNA microarray was scanned for the fluorescent signals and the display of differences between the 2 tissues. IGF1 mRNA, one of the up regulated genes was detected by in situ hybridization technique in the two tissues to validate the result from cDNA microarray.</p><p><b>RESULTS</b>The result indicated that the expressions of 263 genes were up regulated while the expressions of 94 genes were down regulated in the polycystic kidney tissue among the 8398 target genes. Bioinformatical analysis of those genes had been performed. The up-regulated genes were mainly the ones of oncogene, cellular skeleton and movement, apoptosis related protein, cell signal transduction protein, and cytokine. The down regulated genes were mainly the ones of anti-oncogene, DNA binding and transcription factors, cell signal transduction protein, and metabolism protein. The IGF1 mRNA expression detected by in situ hybridization was consequently consistent with the cDNA microarray.</p><p><b>CONCLUSION</b>cDNA microarray is an effective and quick method for studying differential expressed genes. Three hundred and fifty-seven differentially expressed genes with different functions were revealed in the polycystic kidney tissue, which may play some roles in the progression of polycystic kidney.</p>

Expression of PKD1 and PKD2 transcripts and proteins and its significance in different types of kidney tissues and kidney lines / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the expression and function of PKD1 and PKD2 in different kidney tissues and cell lines.</p><p><b>METHODS</b>Immunoprecipitation, Western blotting, In situ hybridization and immunohistochemical staining methods were used to observe the expression of PKD1 mRNA and PKD2 mRNA and their protein abundance in different kidney tissues and cell lines.</p><p><b>RESULTS</b>Coordinate expressions of PKD1 and PKD2 were found in all kidney tissues and cell lines. Distribution of PKD1 mRNA and PKD2 mRNA and their protein polycystin-1 and polycystin-2 in normal human adult kidney tissue were mainly expressed in the medullary collecting ducts and distal tubules. Positive staining was also found in the majority of cyst-lining epithelial cells of PKD1 cystic kidney tissue, PKD1 cyst-lining epithelia cell line and LLC-PK1. The expression level of them in cystic epithelia of ADPKD kidney tissue was much higher than that in adult renal tubules (P < 0.01).</p><p><b>CONCLUSIONS</b>Similar expression pattern of PKD1 and PKD2 and their different tissue distribution in different kidney tissues show that the molecular mutuality of PC-1 and PC-2 might be the base of their functional correlation. Polycystins might play an important role in the maintenance of tubular architecture.</p>

Gene diagnosis of autosomal dominant polycystic kidney disease type 2 using microsatellite DNA tightly linked to polycystic kidney disease gene 2 / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To use microsatellite DNA tightly linked to polycystic kidney disease gene 2 in the gene diagnosis of autosomal dominant polycystic kidney disease type 2.</p><p><b>METHODS</b>Microsatellite DNA of D4S1534, D4S1542, D4S1563,D4S2460 and D4S423 were amplified with PCR and the fragments of products were analyzed by capillary electrophoresis and Genescan and Genotyper software, and then gene diagnosis of the pedigrees was made by linkage analysis.</p><p><b>RESULTS</b>Three families were found to be linked to PKD2 in 20 families. Two carriers of PKD2 mutation were revealed by linkage analysis.</p><p><b>CONCLUSION</b>Gene diagnosis can be done for PKD2 mutation carriers prior to cytogenesis. Linkage analysis is a rapid, simple method for studying the heterogeneity of polycystic kidney disease and for diagnosing the disease at the molecular level.</p>