Experimental Study on the Mechanism of Mangiferin Inhibiting Malignant Biological Characteristics of Multiple Myeloma and Exerting Anticancer Effect / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of pure Chinese herbal extract Mangiferin on the malignant biological behaviors of multiple myeloma (MM) cells, and to analyze the molecular mechanism of the anti-myeloma effect of Mangiferin, so as to provide experimental basis for MM replacement therapy.@*METHODS@#U266 and RPMI8226 of human MM cell lines were intervened with different concentrations of Mangiferin. Cell proliferation was detected by CCK-8 method. Annexin V/PI double staining flow cytometry was used to detect cell apoptosis. Western blot was used to detect the expression of apoptosis and related signaling pathway proteins, and real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of matrix metalloproteinase (MMP) and CXC chemokine receptor (CXCR) family.@*RESULTS@#Mangiferin could inhibit the proliferation activity of U266 and RPMI8226 cells and induce cells apoptosis. After Mangiferin intervened in U266, RPMI8226 cells for 48 h, the expression of Bcl-2 family pro-apoptotic protein Bax was up-regulated, while the expression of survivin and Bcl-xL proteins was down-regulated and caspase-3 was hydrolyzed and activated to promote cell apoptosis, besides, the expression of Bcl-2 protein in U266 cells was also significantly down-regulated to induce apoptosis (P<0.05). After Mangiferin intervenes in MM cells, it can not only increase the expression level of tumor suppressor p53, but also induce programmed cell death of MM cells by inhibiting the expression of anti-apoptotic molecules and down-regulating the phosphorylation levels of AKT and NF-κB. In addition, after the intervention of Mangiferin, the expressions of CXCR4, MMP2 and MMP9 in U266 cells were down-regulated (P<0.05), while there is no effect on the expressions of CXCR2, CXCR7 and MMP13 (P>0.05). However, the expressions of CXCR4, MMP9, and MMP13 in RPMI8226 cells were down-regulated (P<0.01), the expression of MMP2 was weakly affected, and the expression of CXCR2 and CXCR7 was basically not affected (P>0.05).@*CONCLUSION@#Mangiferin can inhibit the proliferation and induce apoptosis of MM cells, and its mechanism may be related to inhibiting the activation of NF-κB signaling pathway, affecting the expression of Bcl-2 family proteins, and inhibiting the expression of core members of MMP and CXCR family.

The Effects and Regulatory Mechanism of Targeting CXC Chemokine Receptor 1/2 Combined with Ara-C on the Malignant Biological Behaviors of U937 Cells of Acute Myeloid Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate and analyze the effect of CXC chemokine receptor 1/2 (CXCR1/2) targeting inhibitor Reparixin combined with cytarabine (Ara-C) on the malignant biological behaviors of acute myeloid leukemia cells and its effect on the expression of the CXCR family, while exploring the accompanying molecular mechanism, providing scientific basis and reference for new molecular markers and targeted therapy for AML.@*METHODS@#Acute myeloid leukemia U937 cells were treated with different concentrations of Reparixin, Ara-C alone or in combination, and the cell morphology was observed under an inverted microscope; Wright-Giemsa staining was used to detect cell morphological changes; CCK-8 method was used to detect cell proliferation; the ability of cell invasion was detected by Transwell chamber method; the ability of colony formation was detected by colony formation assay; cell apoptosis was detected by Hoechst 33258 fluorescent staining and Annexin V/PI double-staining flow cytometry; monodansylcadaverine(MDC) staining was used to detect cell autophagy; the expression of apoptosis, autophagy and related signaling pathway proteins was detected by Western blot and the expression changes of CXCR family were detected by real-time quantitative polymerase chain reaction (qRT-PCR).@*RESULTS@#Reparixin could inhibit the proliferation, invasion, migration and clone formation ability of U937 cells. Compared with the single drug group, when U937 cells were intervened by Reparixin combined with Ara-C, the malignant biological behaviors such as proliferation, invasion and colony formation were significantly decreased, and the levels of apoptosis and autophagy were significantly increased (P<0.01). After Reparixin combined with Ara-C intervenes in U937 cells, it can up-regulate the expression of the pro-apoptotic protein Bax and significantly down-regulate the expression of the anti-apoptotic protein Bcl-2, and also hydrolyze and activate Caspase-3, thereby inducing cell apoptosis. Reparixin combined with Ara-C could up-regulate the expressions of LC3Ⅱ and Beclin-1 proteins in U937 cells, and the ratio of LC3Ⅱ/LC3Ⅰ in cells was significantly up-regulated compared with single drug or control group (P<0.01). MDC result showed that the green granules of vesicles increased significantly, and a large number of broken cells were seen (P<0.01). Reparixin combined with Ara-C can significantly inhibit the phosphorylation level of PI3K, AKT and NF-κB signaling molecule, inhibit the malignant biological behavior of cells by inhibiting the activation of PI3K/AKT/NF-κB pathway, and induce programmed cell death. Ara-C intervention in U937 cells had no effect on the expression of CXCR family (P>0.05). The expression of CXCR1, CXCR2, and CXCR4 mRNA could be down-regulated by Reparixin single-agent intervention in U937 cells (P<0.05), and the expression of CXCR2 was more significantly down-regulated than the control group and other CXCRs (P<0.01). When Reparixin and Ara-C intervened in combination, the down-regulated levels of CXCR1 and CXCR2 were more significant than those in the single-drug group (P<0.01), while the relative expressions of CXCR4 and CXCR7 mRNA had no significant difference compared with the single-drug group (P>0.05).@*CONCLUSION@#Reparixin combined with Ara-C can synergistically inhibit the malignant biological behaviors of U937 cells such as proliferation, invasion, migration and clone formation, and induce autophagy and apoptosis. The mechanism may be related to affecting the proteins expression of Bcl-2 family and down-regulating the proteins expression of CXCR family, while inhibiting the PI3K/AKT/NF-κB signaling pathway.

Pilot study of artificial intelligence ultrasound diagnosis of biliary atresia based on deep learning / 中华超声影像学杂志

Objective:To explore the feasibility of artificial intelligence ultrasound to diagnose of biliary atresia (BA) based on deep learning.Methods:A total of 531 gallbladder ultrasound images in 177 cases of BA patients (BA group) and 585 gallbladder ultrasound images in 195 cases of Non-BA patients (Non-BA group) were collected in Hunan Children′s Hospital from September 2018 to October 2020. For the BA and Non-BA groups, all images were divided into training set and test set according to the ratio of 2∶1. The Mask R-CNN model was trained by training samples, and then the model was tested, according to patient and image as a unit respectively, to evaluate the gallbladder organ detection rate and the diagnostic accuracy of BA. In addition, the images of the test set were randomly numbered.Four sonographers were invited to interpret the images to calculate the diagnostic accuracy individually. Last, the diagnostic accuracy of the Mask R-CNN model was compared with that of sonographers.Results:In terms of the automatic detection of gallbladder organs, the detection rate in both BA and Non-BA group reached 100%, but there were 17 false alarms in 372 test images, with a false alarm rate of 4.57%. In terms of the diagnostic rate of gallbladders, when taking patient as a unit, the total diagnostic accuracy of the model in the test set was 95.97%, which was higher than that of the sonographers in other hospitals and the sonographer with intermediate professional title in our hospital, and the difference was statistically significant ( P<0.005). It was slightly higher than that of sonographer with senior professional title in our hospital (91.94%), but the difference was not statistically significant ( P=0.183). When taking picture as a unit, the total diagnostic accuracy of the model was 97.04%, which was higher than that of the sonographers in other hospitals and the sonographer with intermediate professional title in our hospital, and the difference was statistically significant ( P<0.001). It was slightly higher than that of sonographer with senior professional title in our hospital (94.09%), but the difference was not statistically significant ( P=0.05). Conclusions:The artificial intelligence technology based on Mask R-CNN can automatically and accurately detect gallbladder organs and diagnose BA, which is worthy of further study.

Hyperbaric oxygen treatment on keloid tumor immune gene expression / 中华医学杂志(英文版)

BACKGROUND@#Hyperbaric oxygen treatment (HBOT) has been demonstrated to influence the keloid recurrence rate after surgery and to relieve keloid symptoms and other pathological processes in keloids. To explore the mechanism of the effect of HBOT on keloids, tumor immune gene expression and immune cell infiltration were studied in this work.@*METHODS@#From February 2021 to April 2021, HBOT was carried out on keloid patients four times before surgery. Keloid tissue samples were collected and divided into an HBOT group (keloid with HBOT before surgery [HK] group, n = 6) and a non-HBOT group (K group, n = 6). Tumor gene expression was analyzed with an Oncomine Immune Response Research Assay kit. Data were mined with R package. The differentially expressed genes between the groups were compared. Hub genes between the groups were determined and verified with Quantitative Real-time PCR. Immune cell infiltration was analyzed based on CIBERSORT deconvolution algorithm analysis of gene expression and verified with immunohistochemistry (IHC).@*RESULTS@#Inflammatory cell infiltration was reduced in the HK group. There were 178 upregulated genes and 217 downregulated genes. Ten hub genes were identified, including Integrin Subunit Alpha M (ITGAM), interleukin (IL)-4, IL-6, IL-2, Protein Tyrosine Phosphatase Receptor Type C (PTPRC), CD86, transforming growth factor (TGF), CD80, CTLA4, and IL-10. CD80, ITGAM, IL-4, and PTPRC with significantly downregulated expression were identified. IL-10 and IL-2 were upregulated in the HK group but without a significant difference. Infiltration differences of CD8 lymphocyte T cells, CD4 lymphocyte T-activated memory cells, and dendritic resting cells were identified with gene CIBERSORT deconvolution algorithm analysis. Infiltration levels of CD4 lymphocyte T cell in the HK group were significantly higher than those of the K group in IHC verification.@*CONCLUSION@#HBOT affected tumor gene expression and immune cell infiltration in keloids. CD4 lymphocyte T cell, especially activated memory CD4+T, might be the key regulatory immune cell, and its related gene expression needs further study.

MicroRNA-663 facilitates the growth, migration and invasion of ovarian cancer cell by inhibiting TUSC2

BACKGROUND: MicroRNAs (miRNAs) have emerged as the critical modulators of the tumorigenesis and tumor progression. METHODS: The levels of miR-663 in ovarian cancer cell lines and clinical tissues were detected using qRT-PCR assays. The Transwell invasion and wound healing assay were conducted to assess the roles of miR-663 in the migration and invasion of ovarian cancer cell in vitro. Rescue assays were carried out to confirm the contribution of tumor suppressor candidate 2 (TUSC2) in the aggressiveness of cancer cell which was regulated by miR-663. RESULTS: The levels of miR-663 were up-regulated in ovarian cancer tissues in comparison with the corresponding normal tissues. Up-regulation of miR-663 increased the proliferation, colony formation, migration and invasion of ovarian cancer SKOV3 cell. Additional, over-expression of miR-663 increased the tumor growth of SKOV3 in xenograft model. Bioinformatics analysis and luciferase reporter assay identified that miR-663 decreased the level of TUSC2 via binding to the 3'-UTR of TUSC2 gene. Finally, the expression of TUSC2 was inversely associated with the level of miR-663 in ovarian carcinoma tissue and over-expression of TUSC2 inhibited the migration and invasion abilities of SKOV3 that was promoted by miR-663. CONCLUSION: Altogether, these results indicate that miR-663 acts as a potential tumor-promoting miRNA through targeting TUSC2 in ovarian cancer.

Ki-67 expression affects the clinical therapeutic effect on advanced lung adenocarcinoma / 医学研究生学报

Objective The expression level of the Ki-67 protein is a marker of the poor prognosis in non-small-cell lung cancer. This study explored the correlation of the Ki-67 expression level with the clinical efficacy in the treatment of advanced lung adenocarcinoma.Methods From January 2015 to December 2015, 92 patients with stage Ⅳ lung adenocarcinoma were treated in the Department of Respiratory and Critical Care Medicine of Nanjing General Hospital, 65 with oral gefitinib for EGFR mutation-positive or crizotinib for ALK positive (the targeted therapy \[TT\] group) and the other 27 with pemetrexed + cisplatin or nedaplatin (the chemotherapy \[CT\] group). The expression of Ki-67 was determined by immunohistochemistry, its relationship with the clinicopathological features and therapeutic effects was analyzed, and the progression-free survival (PFS) was calculated.Results Ki-67 was expressed lowly in 43 (46.7%) and highly in 49 (53.3%) of the cases. The median PFS after treatment was significantly longer in the patients with a low than in those with a high Ki-67 expression in the TT group (10 vs 7 mo, P<0.05) and in the CT group as well (8 vs 7 mo, P<0.05). Ki-67 (HR=1.011, 95% CI: 1.000-1.023) and ALK (HR=0.325, 95% CI: 0.112-0.942) were shown to be predictive factors of poor PFS.Conclusion The expression level of Ki-67 affects the effect of clinical treatment, especially that of the first-line targeted therapy, on advanced lung adenocarcinoma.

In vivo Breath Analysis by Extractive Electrospray Ionization-Mass Spectrometry for Investigation of Metabolic Responses to Traditional Chinese Medicine Massages / 分析化学

Traditional Chinese Medicine massage is a kind of physiotherapy which affects on specific parts of the body surface by means of training to regulate the function of the body to achieve the therapeutic effect. In this work,under positive detection model, the chemical fingerprint of exhaled breath from volunteers before and after receiving Traditional Chinese Medicine massage within m/z 50-1000 were detected by extractive electrospray ionization-mass spectrometry (EESI-MS). And through high resolution mass spectrometry analysis, the metabolites such as epinephrine (m/z 184. 0889), 3-(3-hydroxyphenyl) propionic acid (m/z 167.0615) and L-tryptophan (m/z 205. 0933) were successfully identified. Besides, chemical fingerprints of volunteers before and after receiving Traditional Chinese Medicine massage under different health condition were clearly differentiated via partial least squares discrimination analysis (PLS-DA). The results showed that Traditional Chinese Medicine massage could significantly change the metabolic process of volunteers. Moreover, it further indicated that the established method could provide a real time fashion to follow metabolic changes caused by Traditional Chinese Medicine massage.

Application of automated breast volume scanner and mammography in differentiation of small breast lesions with BI-RADS-US 4 / 中南大学学报(医学版)

To evaluate the value of automatic breast volume scanner (ABVS) and mammography (MG) in differential diagnosis for small breast lesions with breast imaging reporting and data system ultrasound (BI-RADS-US) 4.
 Methods: ABVS and MG were performed for 103 patients with 109 breast lesions, which were classified as BI-RADS-US 4 by conventional ultrasound (US). Postoperative pathological diagnosis served as gold standard. The diagnostic efficacy for US, US combined with MG, US combined with ABVS and the combination of three methods were compared.
 Results: The sensitivity of US, US combined with MG, US combined with ABVS and the combination of three methods were 85.5%, 86.8%, 94.7% and 96.0%, respectively. The specificity for them were 66.7%, 69.7%, 81.8% and 84.9%, respectively. The accuracy for them were 79.8%, 81.6%, 90.8% and 92.7%, respectively. The area under the receiver operating characteristic (ROC) curve (AUC) for them were 0.76, 0.78, 0.88 and 0.90, respectively. The accuracy and AUC for US combined with ABVS in differential diagnosis of BI-RADS-US 4 small breast lesions were significantly higher than those of US and US combined with MG (P0.05). No significant difference was found in sensitivity, specificity, accuracy, and AUC between US combined with ABVS and the combination of three methods (P>0.05).
 Conclusion: Combination of US with ABVS can improve the diagnostic efficacy of conventional US in differential diagnosis for BI-RADS-US 4 small breast lesions, and it is superior to US combined with MG.

Simutaneous determination of ursolic acid and oleanolic acid in Ziziphora clinopodioides Lam. by in situ pretreatment-thin layer chromatography scanning / 国际药学研究杂志

Objective To establish a method for the simultaneous determination of ursolic acid(UA)and oleanolic acid(OA) in Ziziphora clinopodioides Lam.,and the quantitative determination the UA and OA contents in the different Z. clinopodioides plant samples collected with various parts of the plant at different times,from different regions of Xinjiang,China. Methods Dual wave?length scanning method was used for the quantification of UA and OA spots on a silica gel G plate in the TLC analysis. The samples loaded on the silica gel G plate were in situ treated with the 1%iodine solution in dichloromethane,and then the plate was developed using cyclohexane,cyclohexane-chloroform-ethyl acetate-formic acid(20:5:8:0.1)as the developing solvent. In the dual wavelength scanning,the measurement wavelength was 530 nm and the reference wavelength was 700 nm. Results The UA and OA spots in samples were well separated on the TLC plate and could be simultaneously quantified by the present method. The average contents of UA and OA in Z. clinopodioides plant samples from 18 different areas were(1.84 ± 0.41)and(2.82 ± 0.89)mg/g,respectively. The contents of UA and OA in the plant increased from late spring to early summer and then decreased thereafter. As to the different parts of the plant,the contents of UA and OA were highest in leaves and lowest in stems. Conclusion The method is simple,fast and accu?rate. The present results provided basic data for further evaluation of the quality of Z. clinopodioides resources.

Heart Rate Variability in Patients with Acute Ischemic Stroke at Different Stages of Renal Dysfunction: A Cross-sectional Observational Study / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Renal function is associated with mortality and functional disabilities in stroke patients, and impaired autonomic function is common in stroke, but little is known regarding its effects on stroke patients with renal dysfunction. This study sought to evaluate the association between autonomic function and stroke in patients with renal dysfunction.</p><p><b>METHODS</b>This study comprised 232 patients with acute ischemic stroke consecutively enrolled from February 2013 to November 2014 at Guangdong Provincial Hospital of Chinese Medicine in China. All patients recruited underwent laboratory evaluation and 24 h Holter electrocardiography (ECG). Autonomic function was measured based on the heart rate variability (HRV) using 24 h Holter ECG. Renal damage was assessed through the estimated glomerular filtration rate (eGFR), and stroke severity was rated according to the National Institutes of Health Stroke Scale (NIHSS). The Barthel index and modified Rankin score were also determined following admission. All the clinical covariates that could potentially affect autonomic outcome variables were adjusted with linear regression.</p><p><b>RESULTS</b>In the patients with a mild or moderate decreased eGFR, the values for the standard deviation of the averaged normal-to-normal RR interval (SDANN) index (P = 0.022), very low frequency (VLF) (P = 0.043), low frequency (LF) (P = 0.023), and ratio of low-to-high frequency power (LF/HF) (P = 0.001) were significantly lower than those in the patients with a normal eGFR. A multinomial linear regression indicated that eGFR (t = 2.47, P = 0.014), gender (t = -3.60, P < 0.001), and a history of hypertension (t = -2.65, P = 0.008) were the risk factors of LF/HF; the NIHSS score (SDANN index: t = -3.83, P < 0.001; VLF: t = -3.07, P = 0.002; LF: t = -2.79, P = 0.006) and a history of diabetes (SDANN index: t = -3.58, P < 0.001; VLF: t = -2.54, P = 0.012; LF: t = -2.87, P = 0.004) were independent factors for the SDANN index, VLF, and LF; the Oxfordshire Community Stroke Project (t = -2.38, P = 0.018) was related to the SDANN index.</p><p><b>CONCLUSIONS</b>Autonomic dysfunction is aggravated with the progression of eGFR stage in patients with acute ischemic stroke; the eGFR is an independent factor of LF/HF in the adjusted models. Stroke severity and a history of diabetes are more significantly associated with HRV in patients with acute ischemic stroke at different stages of renal dysfunction.</p>

Short-term Preoperative Octreotide for Thyrotropin-secreting Pituitary Adenoma / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Thyrotropin-secreting pituitary adenomas (TSHomas) are a rare cause of hyperthyroidism. Somatostatin (SST) analogs work by interacting with somatostatin receptors (SSTRs). This study aimed to evaluate short-term preoperative octreotide (OCT) use in TSHoma patients and to investigate SSTR2 and SSTR5 expression and observe structural changes in tumor tissue.</p><p><b>METHODS</b>We reviewed records and samples from eight TSHoma patients treated between July 2012 and July 2015. We tested immunohistochemically for SSTR2/5 expression and examined TSHoma cells for morphological changes. Signed rank sum test was used to compare the efficacy of short-term preoperative OCT treatment.</p><p><b>RESULTS</b>OCT treatment (median time: 7.9 days, range: 3-16 days; median total dose: 1.8 mg, range: 0.9-4.2 mg) led to significant decrease in all patients' thyroid hormone levels (FT3 [nmol/L]: 8.33 [7.02, 12.29] to 4.67 [3.52, 5.37] [P = 0.008]; FT4 [pmol/L]: 25.36 [21.34, 28.99] to 16.66 [14.88, 21.49] [P = 0.016]; and TSH [μU/ml]: 5.80 [4.37, 6.78] to 0.57 [0.19, 1.24] [P = 0.008]). All the eight tumor specimens expressed high SSTR2 protein levels; 5/8 expressed high SSTR5, but 3/8 that expressed low SSTR5 presented a significantly higher TSH suppression rate (P = 0.036). Electron microscopy showed subcellular level impairments, including clumped nuclear chromatin and reduced cytoplasmic volume. Golgi complexes were observed in the OCT-treated TSHoma specimens.</p><p><b>CONCLUSIONS</b>OCT can control hormone levels and damage the ultrastructure of tumor cells and organelles. Short-term response to OCT may be related to SSTR5 expression. Preoperative SST analog treatment for TSHoma could be considered as a combination therapy.</p>

Pedicle screw fixation with kyphoplasty decreases the fracture risk of the treated and adjacent non-treated vertebral bodies: a finite element analysis / 华中科技大学学报（医学）（英德文版）

Adjacent vertebral fractures are common in patients with osteoporotic vertebral compression fractures (OVCFs) after kyphoplasty. This finite element study was to examine whether short segment pedicle screw fixation (PSF) with kyphoplasty may decrease the fracture risk of the treated and adjacent non-treated vertebrae after kyphoplasty for OVCFs. By simulating cement augmentation with or without short segment pedicle screw fixation (PSF), two tridimensional, anatomically detailed finite element models of the T10-L2 functional spinal junction were developed. The insertion of pedicle screws into the intact vertebra apparently decreased the stress distribution of the treated vertebra in vertical compression and other load situations. The stress distribution in the bone structures of the intact vertebra adjacent to the intact-screwed vertebra was much less than that in the one adjacent to the treated vertebra. The insertion of pedicle screws into the intact vertebra greatly decreased the maximum displacement of the cortical bones and cancellous bones of the vertebrae. Our results indicated that short segment PSF with kyphoplasty may decrease the fracture risk of the treated and adjacent non-treated vertebrae in the management of OVCFs.

Poly(ADP-ribosyl)ation of Apoptosis Antagonizing Transcription Factor Involved in Hydroquinone-Induced DNA Damage Response / 生物医学与环境科学（英文）

The molecular mechanism of DNA damage induced by hydroquinone (HQ) remains unclear. Poly(ADP-ribose) polymerase-1 (PARP-1) usually works as a DNA damage sensor, and hence, it is possible that PARP-1 is involved in the DNA damage response induced by HQ. In TK6 cells treated with HQ, PARP activity as well as the expression of apoptosis antagonizing transcription factor (AATF), PARP-1, and phosphorylated H2AX (γ-H2AX) were maximum at 0.5 h, 6 h, 3 h, and 3 h, respectively. To explore the detailed mechanisms underlying the prompt DNA repair reaction, the above indicators were investigated in PARP-1-silenced cells. PARP activity and expression of AATF and PARP-1 decreased to 36%, 32%, and 33%, respectively, in the cells; however, γ-H2AX expression increased to 265%. Co-immunoprecipitation (co-IP) assays were employed to determine whether PARP-1 and AATF formed protein complexes. The interaction between these proteins together with the results from IP assays and confocal microscopy indicated that poly(ADP-ribosyl)ation (PARylation) regulated AATF expression. In conclusion, PARP-1 was involved in the DNA damage repair induced by HQ via increasing the accumulation of AATF through PARylation.

Analysis on DNA methylation of mosaic and moxa type leaves of Aconitum carmichaeli by MSAP / 中国中药杂志

An MSAP analysis method was established for detecting DNA methylation of Aconitum carmichaeli leaves, and the DNA methylation of different leaf shapes and different leaf position was analyzed by MSAP. The study made experiments on the leaves of different position of mosaic and moxa leaf type A. carmichaeli, researched the effects of restriction digestion of genomic DNA by using two restriction enzymes, screened the suitable selective amplification primers, and analyzed the methylation differences of leaves by calculating the 6% acrylamide gel electrophoresis bands and lane. The best reaction system of MSAP was obtained, under the conditions of 37 ℃, the 16 h incubated time was more suitable for 150 ng DNA, and 25 pairs of selective amplification primers were selected from 256 pairs. Totally, 273 electrophoresis bands were obtained by 25 pairs of selective primers, including 228 non methylation or single chain methylation bands,27 double chain methylation bands,and 18 single stranded methylation bands, the total methylation rate was 16.48%. The methylation rate was slightly different in mosaic and moxa leaf type A. carmichaeli leaf, which were 15.36%, 14.34%, respectively, and article 8, article 6 nucleotide fragments of genome methylation modification differences were obtained, accounted for 3%, 2.26% of the total number of bands. Based on this study it can provide new ideas for molecular identification, breeding and cultivation, and genetic evolution of A. carmichaeli.

Primary culture and characteristics of colorectal cancer-associated fibroblasts / 中华病理学杂志

<p><b>OBJECTIVE</b>To compare the biological characteristics of colorectal cancer associated fibroblasts (CAFs) with normal fibroblasts (NFs).</p><p><b>METHODS</b>CAFs and NFs were isolated from fresh specimens of colorectal cancer and their paired normal colon tissue and cultured by tissue explant method. Light microscopy, quantitative polymerase chain reaction (qPCR), Western blot, immunofluorescence microscopy, electron microscopy and flow cytometry were used to identify isolated fibroblasts and to explore their characteristics of activation and growth.</p><p><b>RESULTS</b>Primary colorectal CAFs and NFs were isolated and cultured successfully. NFs showed spindled morphology and were arranged in interlacing or spiral bundles. CAFs were polygonal or spindle, but were fatter than NFs. They were distributed randomly and arranged irregularly, and had obvious actin expression. CAFs and NFs both expressed fibronectin, but not E-cadherin, CD31 and caldesmon. qPCR showed that CAFs expressed more fibroblast activation protein (FAP) and less fibroblast specific protein 1 (FSP1) than that of NFs. There was no difference in the expression of α-SMA between NFs and CAFs by Western blot. α-SMA was bundled in parallel to the long axis of the cell by immunofluorescence. By electron microscopy, CAFs but not NFs showed dense myofilament that was arranged regularly. Flow cytometry showed that the percentage of S- and G2-phase in CAFs were significantly lower than that in NFs. mRNA expression of transforming growth factor β1, stromal derived factor 1 (SDF-1) and platelet derived growth factor (PDGF)-D in CAFs were lower while that for PDGFC was higher than that in NFs. That indicated the proliferation of CAFs was inhibited and the secretion of some cytokines was different when compared with NFs.</p><p><b>CONCLUSIONS</b>CAFs show differences with NFs in morphology, characteristics of activation and secretion of some cytokines. The proliferation of CAFs is down regulated as compared with NFs.</p>

Fourier transform infrared spectroscopy in detection of breast cancer / 浙江大学学报·医学版

Breast cancer is the most common cancer in women. Later diagnosis of the disease is the leading cause of poor prognosis. Fourier-transform infrared (FTIR) spectroscopy is a novel approach to provide information about the molecular components and metabolic conditions of human tissue; therefore it can detect the early changes caused by cancer cells prior to histological manifestation. FTIR-based diagnosis is rapid, simple and label free, which meets the requirements of an automated and patient-friendly technique. The current article gives an overview of the experimental techniques, data analysis methods and spectral signatures of breast cancer in FTIR-based diagnosis, summarizes the present challenges by focusing on the history of FTIR spectroscopy in breast cancer since 1990s, and highlights some investigations that give a perspective of FTIR-based diagnosis.

Establishment of a two-layer cell spheroid model and its applications in colorectal tumor cell-fibroblast interactions with effect on Cx43 expression / 中华病理学杂志

<p><b>OBJECTIVE</b>To imitate tumor microenvironment in vivo through construction of two-layer cell spheroid as a three-dimensional tumor model, and to validate its application in the study of Cx43 expression in colorectal cancer cell-fibroblast interactions and colorectal cancer progression.</p><p><b>METHODS</b>The two-layer cell spheroid was constructed from SW620 colorectal cancer cells and HELF fibroblasts. The expression of Cx43 in the spheroid was detected by immunocytochemistry. The expression of Cx43 in cultured SW620 cells with or without co-cultured fibroblasts was detected by immunocytochemistry and immunofluorescence. The expression of Cx43 in colorectal cancer tissue was detected by immunohistochemistry.</p><p><b>RESULTS</b>The spheroid showed well-defined cellular morphology and clear boundary between two cell lines.Significant expression of Cx43 was found along the boundary.SW620 cells had no expression of Cx43 when cultured alone, while the expression of Cx43 was induced upon co-culturing with fibroblasts.In the colorectal cancer tissue, expression of Cx43 was minimal in the centre of tumor in contrast to an upregulated expression at invasive front.</p><p><b>CONCLUSIONS</b>The two-layer cell spheroid is an observable and sensitive model for cell-cell interaction for studies of tumor microenvironment.It can simulate colorectal cancer cell-fibroblast interactions through up-regulation of Cx43 expression.</p>

Crosstalk between canonical TGF-β/Smad and Wnt/β-catenin signaling pathway / 浙江大学学报·医学版

TGF-β signaling pathway plays a central role in the signaling networks that control the growth, differentiation of the cell, and the initiation of fibrosis and cancer. Wnt signaling pathway is critical for the embryonic development and the invasion and migration of cancer cells. TGF-β signaling and Wnt signaling, both of which play an important role in regulating embryonic development, fibrotic disease and tumor progression, have a close relationship. Researches find several typical cross points between these two signaling systems, such as Smad, Axin, Dvl and β-catenin. In this review, we focus on the crosstalk between TGF-β signaling and Wnt signaling through these typical factors, intending to better understand the process of fibrosis and tumor progression.

Clinicopathologic characteristics of primary thyroid-like follicular carcinoma in kidney / 中华病理学杂志

<p><b>OBJECTIVE</b>To explore the clinicopathological characteristics of primary thyroid-like follicular carcinoma of the kidney.</p><p><b>METHODS</b>A case of primary thyroid-like follicular carcinoma of the kidney was studied with histology and immunohistochemical staining, and its clinical and pathological findings were further analyzed with review of the literature.</p><p><b>RESULTS</b>The patient was a 26-year-old asymptomatic woman who had a kidney mass during her annual physical examination. The tumor was well-circumscribed. Pathologically, the tumor showed follicular structures with colloid-like material in the lumina. Immunohistochemically, the tumor cells showed intense staining for CK7 and vimentin and negative for thyoid transcripation factor-1, thyroglobulin, thyoid peroxidase and RCC.</p><p><b>CONCLUSIONS</b>The diagnosis of primary thyroid-like follicular carcinoma of the kidney is based on the characteristic follicular architecture with colloid-like material, and the metastasis from a thyroid follicular carcinoma must be excluded clinically and pathologically before making the final diagnosis.</p>

Effects of different therapeutic methods and typical recipes of Chinese medicine on activation of c-Jun N-terminal kinase in kupffer cells of rats with fatty liver disease / 中国结合医学杂志

<p><b>OBJECTIVE</b>To observe the effects of different therapeutic methods and the recipes of Chinese medicine (CM) on the activation of c-Jun N-terminal kinase (JNK) in Kupffer cells of rats with fatty liver disease and to explore the mechanisms of these therapeutic methods.</p><p><b>METHODS</b>By using a random number table, 98 rats were randomly divided into 7 groups: control group, model group, and 5 treatment groups, including soothing Liver (Gan) recipe group, invigorating Spleen (Pi) recipe group, dispelling dampness recipe group, promoting blood recipe group, and complex recipe group. Rats in the control group were fed with normal food and distilled water by gastric perfusion, while rats in the model group were fed with high-fat food and distilled spirits by gastric perfusion. Rats in the 5 treatment groups were fed with high-fat food and corresponding recipes by gastric perfusion. Twelve weeks later, all rats were sacrificed and liver tissues were stained for pathohistological observation. Kupffer cells were isolated from livers of rats to evaluate JNK and phospho-JNK expressions by Western blotting.</p><p><b>RESULTS</b>The grade of hepatic steatosis was higher in the model group than the control group (P<0.05). Compared with the model group, the grade of fatty degeneration in soothing Liver recipe group and invigorating Spleen recipe group were significantly ameliorated (P<0.05). Expressions of JNK and phospho-JNK in Kupffer cells were significantly higher in the model group than those in the control group (P<0.05, P<0.01). Compared with the model group, expressions of JNK in all treatment groups decreased, especially in invigorating Spleen recipe group and promoting blood recipe group (P<0.05). Compared with the model group, expressions of phospho-JNK in all treatment groups declined significantly (P<0.01), especially in soothing Live recipe group and invigorating Spleen recipe group.</p><p><b>CONCLUSIONS</b>The high expressions of JNK and phospho-JNK in Kupffer cells might play an important role in the pathogenesis of fatty liver disease in rats. The recipes of CM, especially invigorating Spleen recipe and soothing Liver recipe, might protect liver against injury by reducing the total JNK protein content and inhibiting the activation of JNK protein in Kupffer cells of fatty liver model rats, which showed beneficial effects on fatty liver disease.</p>