Long non-coding RNA CASC2 induces apoptosis and autophagy in human colon cancer cells via modulation of TRIM16 expression.

Long non-coding RNAs (LncRNAs) have been shown to be involved in diverse cellular and physiological processes. Recent studies have proved their potential as the prospective therapeutic targets for cancer treatment. Herein, we examined the role of LncRNA CASC2 in human colon cancer. The gene expression analysis showed that LncRNA CASC2 is significantly suppressed in colon cancer tissues and cell lines. The immunohistochemistry also showed considerable increase of the Ki67 in colon cancer tissues suggestive of their aggressiveness. Overexpression of CASC2 inhibited the growth of HT-29 cells. The inhibition of HT-29 growth was due to the induction of apoptosis which was accompanied by upsurge of Bax, depletion of Bcl-2 and activation of caspase-3 cleavage. Electron microscopic analysis showed CASC2 overexpression also induced autophagy in the HT-29 cells which was associated with increase in LC3B II and Beclin 1 expression. Bioinformatic approaches and dual luciferase assay showed that CASC2 controls the TRIM16 via microRNA-214 axis. TRIM16 was found to be overexpressed in all the colon cancer tissues and cell lines. Overexpression of CASC2 caused significant inhibition of TRIM16. Additionally, silencing of TRIM16 resulted in the inhibition of HT-29 cell growth similar to that of CASC2 overexpression. Taken together, CASC2 may prove to be an important therapeutic target for colon cancer treatment.

Hypoxic preconditioning attenuates necroptotic neuronal death induced by global cerebral ischemia via Drp1-dependent signaling pathway mediated by CaMKIIα inactivation in adult rats.

Hypoxic preconditioning (HPC) alleviates the selective and delayed neuronal death in the hippocampal CA1 region induced by transient global cerebral ischemia (tGCI). This type of cell death may include different programmed cell death mechanisms, namely, apoptosis and necroptosis. Although apoptotic signaling is well defined, the mechanisms that underlie neuronal necroptosis are yet to be fully elucidated. In this study, we investigated whether HPC protects neurons from cerebral ischemia-induced necroptosis. We observed that tGCI up-regulated the expression of receptor-interacting protein (RIP) 3 and increased the interaction of RIP1-RIP3 in CA1 at the early stage of reperfusion. The pretreatment with HPC or necrostatin-1 decreased the expression of RIP3 and the formation of RIP1-RIP3 after tGCI. We also found that HPC decreased the expression and the activity of caspase-8 in CA1 after tGCI, and notably, the pretreatment with Z-VAD-FMK, a pan-caspase inhibitor, did not trigger necroptosis but attenuated the tGCI-induced neuronal damage. Furthermore, we demonstrated that HPC decreased the activation of calcium-calmodulin kinase (CaMK) IIα and the interaction of RIP1 and CaMKIIα induced by tGCI. Intriguingly, the pretreatment with a CaMKs inhibitor KN-93 before tGCI resulted in significantly reduced RIP1-3 interaction and tGCI-induced neuronal damage. Finally, we ascertained that HPC prevented the dephosphorylation of dynamin-related protein 1 (Drp1)-Ser637 (serine 637) and inhibited the translocation of Drp1 to mitochondria induced by tGCI. Importantly, the treatment with a Drp1 inhibitor Mdivi-1 or necrostatin-1 before tGCI also abolished Drp1 dephosphorylation at Ser637 and mitochondrial translocation. Taken together, our results highlight that HPC attenuates necroptotic neuronal death induced by tGCI via Drp1-dependent mitochondrial signaling pathways mediated by CaMKIIα inactivation.-Zhan, L., Lu, Z., Zhu, X., Xu, W., Li, L., Li, X., Chen, S., Sun, W., Xu, E. Hypoxic preconditioning attenuates necroptotic neuronal death induced by global cerebral ischemia via Drp1-dependent signaling pathway mediated by CaMKIIα inactivation in adult rats.

Autophagosome maturation mediated by Rab7 contributes to neuroprotection of hypoxic preconditioning against global cerebral ischemia in rats.

Autophagy disruption leads to neuronal damage in hypoxic-ischemic brain injury. Rab7, a member of the Rab GTPase superfamily, has a unique role in the regulation of autophagy. Hypoxic preconditioning (HPC) provides neuroprotection against transient global cerebral ischemia (tGCI). However, the underlying mechanisms remain poorly understood. Thus, the current study explored the potential molecular mechanism of the neuroprotective effect of HPC by investigating how Rab7 mediates autophagosome (AP) maturation after tGCI in adult rats. We found that HPC attenuated AP accumulation in the hippocampal CA1 region after tGCI via restoration of autophagic flux. We also confirmed that this HPC-induced neuroprotection was not caused by the increase in lysosomes or the improvement of lysosomal function after tGCI. Electron microscopic analysis then revealed an increase in autolysosomes in CA1 neurons of HPC rats. Moreover, the inhibition of autophagosome-lysosome fusion by chloroquine significantly aggravated neuronal death in CA1, indicating that AP maturation contributes to HPC-induced neuroprotection against neuronal injury after tGCI. Furthermore, the activation of Rab7 was found to be involved in the neuroprotective effect of AP maturation after HPC. At last, the knockdown of ultraviolet radiation resistance-associated gene (UVRAG) in vivo disrupted the interaction between Vps16 and Rab7, attenuated the activation of Rab7, interrupted autophagic flux, and ultimately abrogated the HPC-induced neuroprotection against tGCI. Our results indicated that AP maturation was enhanced by the activation of Rab7 via UVRAG-Vps16 interaction, which further demonstrated the potential neuroprotective role of Rab7 in HPC against tGCI-induced neuronal injury in adult rats.

Repair of Concomitant Incisional and Parastomal Hernias Using a Hybrid Technique: A Series of 32 Patients.

BACKGROUND: Concomitant incisional and parastomal hernias is a challenging condition. We used a hybrid technique of sublay and onlay to treat patients with this condition. MATERIAL AND METHODS: The clinical data of 32 consecutive patients treated from February 2008 to April 2014 for concomitant incisional and parastomal hernias were retrospectively reviewed. The mean diameter was 9 (range 4-13) cm of the incisional hernias, and 6 (range 4.5-8) cm of the parastomal hernias. RESULTS: The mean operative time was 247 (range 220-290) min. The mean hospital stay was 20 (range 14-27) days. All surgical wounds healed by primary intention. Seven patients had postoperative seroma and were well managed with puncture and compression. All 32 patients were followed up for a mean of 48 (range 5-68) months. Four patients recurred with parastomal hernias and were treated with secondary surgery. No further recurrence occurred until the last follow-up. CONCLUSIONS: This hybrid technique of sublay and onlay is only suitable for the repair of complex incisional and parastomal hernias.

Laparoscopic-assisted subtotal colectomy combined with modified Duhamel procedure for mixed constipation.

BACKGROUND AND OBJECTIVES: To evaluate the effects of subtotal colectomy combined with the modified Duhamel procedure on mixed constipation. METHODS: A total of 16 female patients with mixed constipation were enrolled and underwent subtotal colectomy combined with the modified Duhamel procedure under laparoscopy from April 2010 to April 2012. Before surgery, physical examinations such as the gastrointestinal transit test, barium enema, and defecography were performed for all the patients. After surgical treatment, 2-year follow-up was performed using questionnaires to assess the effect of treatment. RESULTS: All 16 cases were treated successfully, with a mean operation time of 230 minutes (range, 180-290 minutes). No intraoperative or postoperative complications were found, and no deaths occurred. Constipation and relevant symptoms were relieved, and all patients were satisfied with their quality of life. The gastrointestinal quality-of-life score was significantly increased 6 months postoperatively (mean, 102) compared with preoperatively (mean, 75). CONCLUSION: Subtotal colectomy combined with the modified Duhamel procedure under laparoscopy is effective and safe for the treatment of mixed constipation.

Microarray technology to investigate genes associated with papillary thyroid carcinoma.

DNA microarray data on thyroid tissue from patients with papillary thyroid carcinoma (PTC) and from healthy controls were compared in order to investigate the regulatory genes and uncover the underlying regulatory network in PTC. The DNA microarray data set, GSE3678, was downloaded from Gene Expression Omnibus database. This included seven thyroid tissue samples from patients with PTC and seven samples from healthy controls. Raw data were processed and differentially expressed genes (DEGs) were identified using corresponding R packages. Gene regulation analysis was conducted using TRANSFAC® and TRED. A total of 171 DEGs were obtained. A regulatory network was then established, using 104 of the DEGs. Subsequently, pathway enrichment analyses of the genes were conducted using Database for Annotation, Visualization and Integrated Discovery (DAVID) online tool. Three differentially expressed transcription factors were identified: Trefoil factor 3, cut­like homeobox 2 and forkhead box protein A2. The most significant pathways involving the 104 DEGs were pathways involved in cancer. Biological process analysis using DAVID, suggested that these genes were associated with the positive regulation of gene expression, gene transcription and metabolic processes. The present study identified a range of genes associated with the development of PTC. The results of the present study were beneficial for understanding the regulatory mechanisms involved in PTC, and for developing clinical diagnostic and therapeutic approaches for this disease.

Hypoxia-inducible factor 1α mediates neuroprotection of hypoxic postconditioning against global cerebral ischemia.

Hypoxia administered after transient global cerebral ischemia (tGCI) has been shown to induce neuroprotection in adult rats, but the underlying mechanisms for this protection are unclear. Here, we tested the hypothesis that hypoxic postconditioning (HPC) induces neuroprotection through upregulation of hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF), and that this involves phosphatidylinositol-3-kinase (PI3K), p38 mitogen-activated protein kinase (p38 MAPK), and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK) pathways. The expression of HIF-1α, VEGF, and cleaved caspase-9 were determined by immunohistochemistry and Western blot. As pharmacologic interventions, the HIF-1α inhibitor 2-methoxyestradiol (2ME2), PI3K inhibitor LY294002, p38 MAPK inhibitor SB203580, and MEK inhibitor U0126 were administered before HPC or after tGCI. We found that HPC maintained the higher expression of HIF-1α and VEGF and decreased cleaved caspase-9 levels in CA1 after tGCI. These effects were reversed by 2ME2 administered before HPC, and the neuroprotection of HPC was abolished. LY294002 and SB203580 decreased the expression of HIF-1α and VEGF after HPC, whereas U0126 increased HIF-1α and VEGF after tGCI. These findings suggested that HIF-1α exerts neuroprotection induced by HPC against tGCI through VEGF upregulation and cleaved caspase-9 downregulation, and that the PI3K, p38 MAPK, and MEK pathways are involved in the regulation of HIF-1α and VEGF.

Identification of collaboration patterns of dysfunctional pathways in breast cancer.

Breast cancer (BC) is the most common malignancy among women. We aimed to illuminate the molecular dysfunctional mechanisms of BC progression. The mRNA expression profile of BC GSE15852 was downloaded from Gene Expression Omnibus database, including 43 normal samples and 43 cancer samples. Differentially expressed genes (DEGs) in BC were screened using the t-test by Benjamin and Hochberg method. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of the selected DEGs were enriched using Hypergeomeric distribution model. In addition, functional similarity network among the enriched pathways was constructed to further analyze the collaboration of these pathways. We found 848 down-regulated DEGs were associated with 16 significant dysfunctional pathways, including PPAR signaling fatty acid metabolism, and 1584 up-regulated DEGs were related to 6 significant dysfunctional pathways, like cell cycle, protein export, and antigen processing and presentation in BC samples. Crosstalk network analysis of pathways indicated that pyruvate metabolism, propanoate metabolism, and glycolysis gluconeogenesis were the pathways with closest connections with other pathways in BC. In addition, other antigen processing and presentation, including 19 DEGs; PPAR signaling pathway, including 18 DEGs; and pyruvate metabolism pathway, including 13 DEGs were further analyzed. Our results suggested that dysfunctional of significant pathways can greatly affect the progression of BC. Several significant disorder pathways were enriched in our comprehensive study. They may provide guidelines to explore the dysfunctional mechanism of BC progression.

[Study on the three dimensional hepatic virtual operation based on the data of 64-slice helical CT scanning].

OBJECTIVE: To study the surgery plan and simulation effect of the three dimensional (3D) hepatic virtual operation based on the data of 64-slice helical CT scanning and to probe the feasibility of the virtual operation based on the FreeForm Modeling System. METHODS: The volunteer liver was scanned to collect two dimensional (2D) DICOM data of 64-slice helical CT scanning and the 3D hepatic and intrahepatic vessels model were reconstructed by MIMICS software. The reconstructed liver, the intrahepatic vessels model and the artificial tumor models were output into the FreeForm Modeling System in the STL format. The device PHANTOM with the characterization of dynamo-feedback was applied to make the operation on the 3D hepatic. RESULTS: The spatial relationship between the tumour and the intrahepatic vessels were clearly observed by rotation and enlargement of the target. According to the operation principle, the left lobe of liver resection was simulated by manipulating the device PHANToM. Through the liver transparence surface, the intrahepatic vessels were easily distinguished. The operation procedure was accord with the clinic hepatic surgery. Meanwhile, during the operation, by adjusting the incision objective intensity, the dynamo-feedback intensity was definitely touched. CONCLUSIONS: By using the FreeForm Modeling System,the hepatic operation procedure can be simulated ahead of time. The operation complication in the practical surgery can be anticipated and the individualization operation schema can be reasonable instituted.

[Image segmentation and three-dimensional reconstruction of the liver based on 64-slice spiral CT scanning data].

OBJECTIVE: To study the segmentation methods of the liver CT images and the value of 3-dimensional (3D) reconstruction of the liver in the planning of hepatic surgery. METHODS: The 2D Digital Imaging and Communications in Medicine (DICOM) format data of the liver obtained from healthy volunteers were transformed into bmp format image, and the liver image segmentation was performed using Photoshop software. The 3D model was reconstructed using MIMICS software. RESULTS: The DICOM format data of the liver obtained by 64 slice spiral CT included totally 658 slice images. The segmented liver image showed clear profiles and complete intrahepatic duct data were reserved. The segmented liver images were free of discontinuation during continuous observation. The liver surface and internal ductal system, including the hepatic arteries and veins, and the hepatic portal system and their branches, were represented clearly. The reconstructed liver allowed clear identification of the anatomic landmark and matched the actual liver volume. The reconstructed ductal structure were distinct and continuous with natural texture. The reconstructed liver and the hepatic internal duct system were simultaneously displayed by adjusting the transparency of the liver, and the blood vessels were also represented. CONCLUSION: Segmentation of the liver images in different phases using Photoshop can be feasible for liver reconstruction. The reconstructed liver and the intrahepatic ductal structure allow vivid 3D observation of the spatial relationship among the major tracts and accurate estimation of the liver volume.

[Expressions and role of endogenetic matrix metalloproteinases-9 and transforming growth factor-beta during wound healing of blast injury].

OBJECTIVE: To investigate the expression of endogenetic matrix metalloproteinases-9 (MMP-9) and transforming growth factor-beta(TGF-beta) and their role in the wound healing of blast injury. METHODS: Rat models of blast injury under a humid and hot environment were established and the effusion from the wound surface was collected at 4, 24, 48 h and 5, 7, 14, 21 and 28 days after injury, respectively. The contents of MMP-9 and TGF-beta in the effusion of the wound were measured by zymography and enzyme-linked immunosorbent assay (ELISA), respectively. RESULTS: During the wound healing of blast injury, MMP-9 and TGF-beta exhibited changes that followed a regular pattern, both reaching the peak value at 48 h after the injury. TGF-beta content reached the another peak on day 7. TGF-beta value and MMP-9 contents decreased in the second week after injury and their reduction was no longer parallel. Administration of tissue inhibitor of metalloproteinase (TIMP) in the early phase of injury showed no obvious effect, but during the 2 weeks after the injury, its administration caused decrease in MMP-9 content and increase in TGF-beta content in the effusion. CONCLUSIONS: In the early phase of wound healing, the elevation of MMP-9 and TGF-beta accelerated cell migration to promote the clearance of the inflammatory necrosis tissues, which might be one of the wound healing mechanisms. But overexpression of MMP-9 in the wound may hinder wound healing, and appropriate use of TIMP can accelerate the delayed wound healing.

Distribution of oval cells and c-myc mRNA expression in mouse hepatocarcinogenesis.

BACKGROUND: This study was designed to assess the roles of oval cells and c-myc mRNA in the process of hepatocarcinogenesis and to clarify the function of carcinogene c-myc in the development of hepatocellular carcinoma (HCC) and the mechanism of inhibitory function of uscharidin on HCC in mouse hepatocarcinogenesis. METHODS: A total of 120 clean SD mice were divided into normal group, cancer induction group, and intervention group. The normal group was fed with standard forage while the rest two groups were given p-dimethylaminoazobenzene (DAB) to induce cancer. Thirteen weeks after induction of cancer, the two groups were fed with standard forage and water. Once the pattern was set up, the intervention group was given uscharidin injection into the abdominal cavity from the first week to the 14th week. On the 2nd, 4th, 6th, 8th, 10th, 12th, 14th, 16th, 18th, 20th, 22nd, and 24th week, all mice were killed and biopsied from the liver lobe for pathological analysis. At the same time, the number of tumor nodes was counted and the expression of c-myc mRNA was tested by RT-PCR. RESULTS: Since the 2nd week after cancer induction, proliferated oval cells could be seen in the portal area. Initially, the oval cells appeared in the cortical layer of the portal area, then proliferated gradually and immigrated into the liver parenchyma. In the period of fibrosis after liver proliferation, proliferated heaps of oval cells were noted in both portal and peripheral areas. In the period of carcinomatous change, oval cells could be seen both outside and inside of cancer nodes, but most of them were distributed outside. The c-myc gene was expressed negatively in the liver tissue of mice. The quantity of the expression began to increase at the time of infection of the liver and tended to increase with the degree of hepatic injury. In the period of canceration, the expression level of c-myc mRNA increased gradually. The intervention of uscharidin could not inhibit but delay the increase of the expression of c-myc mRNA. CONCLUSION: Oval cells are closely related to hepatocarcinoma cells, which play an important role in the occurrence and development of hepatocarcinogenesis. Uscharidin can inhibit the occurrence of hepatocarcinogenesis or local spreading at the early stage of cancer induction by DAB, but it cannot inhibit the expression of c-myc.

The expression of c-kit and proliferating cell nuclear antigen in oval cells of rats with hepatocellular carcinoma.

OBJECTIVE: To study the relationship between oval cells and primary hepatocarcinoma and the expression of c-kit and proliferating cell nuclear antigen (PCNA) in oval cells of rats with hepatocellular carcinoma. METHODS: A hundred and twenty clean SD rats were divided into three groups: normal group, cancer-induction group and intervention group. The normal group was fed with standard forage while the rest two groups were fed with 3'-methyl-2-methylamino-azobenzene (DAB) to induce carcinoma for 14 weeks and then fed with standard forage and water. Uscharidin was injected abdominally to the intervention group from the first week to the 14th week. All rats were killed and biopsy specimens were taken from the left and right liver lobes for immunohistochemical staining of c-kit and PCNA on the 2nd, 4th, 6th, 8th, 10th, 12th, 14th, 16th, 18th, 20th, 22nd, and 24th week. RESULTS: From the 2nd to 14th week after liver infection, c-kit positive cells, mainly oval cells were found in the portal area in the carcinoma-induction group and dotted positive pigmentations in liver lobules. In the 22nd week, a large number of cancerous nodes occurred and nuclei heteromorphism was apparent; the number of positive cell decreased but positive cells could be sparsely observed in cancerous nodes. In the 2nd week of the carcinoma-induction process, PCNA positive cells were oval cells in the portal area. In the 4th week, a lot of hepatic cells were positively stained, especially in the central vein area. In the 6th week, PCNA positive cells could be seen in the lobules of the liver. In the 8th week, the number of PCNA cells decreased comparatively. From the 10th to 14th week, oval cells in the portal area were still over-expressed. From the 16th to 24th week, a large number of cancerous nodes occurred and PCNA was over-expressed in some of them. In necrotic cancerous nodes, the para-cancerous PCNA positive cells were sparsely distributed and their number was less than that of PCNA positive cells of cancerous tissues. CONCLUSIONS: Hepatic stem cells originating from the terminal biliary plexus of the portal area are involved in the development of hepatocarcinoma because c-kit positive cells expressed in cancerous nodes, accompany the whole process of the development. In the middle inflammatory period of carcinoma-induction, the expression of PCNA in hepatic cells peaked, but the index decreased in the late inflammatory period and in the proliferated fibrosis stage. The expression of PCNA is a tortuous process, going up, down, then up again from normal tissues to cancerous tissues. Combined with pathological findings, PCNA can be considered as a warning index for carcinomatous cells.