Heparanase inhibitor OGT 2115 induces prostate cancer cell apoptosis via the downregulation of MCL­1.

Heparanase (HPSE), an endo-ß-D-glucuronidase, cleaves heparan sulfate and serves an important role in the tumor microenvironment and thus in tumorigenesis. HPSE is known to promote tumor cell evasion of apoptosis. However, the underlying mechanism of this requires further study. In the present study, the results demonstrated that myeloid cell leukemia-1 (MCL-1), an antiapoptotic protein, and HPSE were upregulated in prostate cancer tissues compared with adjacent normal tissues. In addition, the HPSE inhibitor, OGT 2115, inhibited PC-3 and DU-145 prostate cancer cell viability in a dose-dependent manner, with IC50 values of 20.2 and 97.2 µM, respectively. Furthermore, annexin V/PI double-staining assays demonstrated that OGT 2115 induced apoptosis in prostate cancer cells. OGT 2115 treatment markedly decreased MCL-1 protein expression levels, whereas RNA interference-mediated downregulation of MCL-1 and OGT 2115 drug treatment synergistically induced apoptosis in PC-3 and DU-145 cells. In vivo, OGT 2115 40 mg/kg (ig) significantly inhibited PC-3 cell xenograft growth in nude mice and increased the positive TUNEL staining rate of xenograft tissues. It was therefore hypothesized that MCL-1 was an important signaling molecule in OGT 2115-induced apoptosis. The results of the present study also demonstrated that the proteasome inhibitor, MG-132, markedly inhibited the downregulation of MCL-1 protein expression levels induced by OGT 2115. However, the protein synthesis inhibitor, cycloheximide, did not affect the role of OGT 2115 in regulating MCL-1. In summary, the results of the present study demonstrated that the proapoptotic activity of OGT 2115 was achieved by downregulating MCL-1.

Olink proteomics profiling platform reveals non-invasive inflammatory related protein biomarkers in autism spectrum disorder.

Background: Owing to the lack of valid biomarkers, the diagnosis of autism spectrum disorder (ASD) diagnosis relies solely on the behavioral phenotypes of children. Several researchers have suggested an association between ASD and inflammation; however, the complex relationship between the two is unelucidated to date. Therefore, the current study aims to comprehensively identify novel circulating ASD inflammatory biomarkers. Methods: Olink proteomics was applied to compare the plasma inflammation-related protein changes in a group of the healthy children (HC, n = 33) and another with ASD (n = 31). The areas under the receiver operating characteristic curves (AUCs) of the differentially expressed proteins (DEPs) were calculated. The functional analysis of the DEPs was performed using Gene Ontology and Kyoto Encyclopedia Genes and Genomes. Pearson correlation tests were used employed to analyze the correlation between the DEPs and clinical features. Results: A total of 13 DEPs were significantly up-regulated in the ASD group compared with the HC group. The four proteins, namely, STAMBP, ST1A1, SIRT2, and MMP-10 demonstrated good diagnostic accuracy with the corresponding AUCs (95% confidence interval, CI) of 0.7218 (0.5946-0.8489), 0.7107 (0.5827-0.8387), 0.7016 (0.5713-0.8319), and 0.7006 (0.568-0.8332). Each panel of STAMBP and any other differential protein demonstrated a better classification performance [AUC values from 0.7147 (0.5858-0.8436, STAMBP/AXIN1) to 0.7681 (0.6496-0.8867, STAMBP/MMP-10)]. These DEP profiles were enriched in immune and inflammatory response pathways, including TNF and NOD-like receptor signaling pathways. The interaction between STAMBP and SIRT2 (R = 0.97, p = 8.52 × 10-39) was found to be the most significant. In addition, several DEPs related to clinical features in patients with ASD, particularly AXIN1 (R = 0.36, p = 0.006), SIRT2 (R = 0.34, p = 0.010) and STAMBP (R = 0.34, p = 0.010), were positively correlated with age and parity, indicating that older age and higher parity may be the inflammation-related clinical factors in ASD. Conclusion: Inflammation plays a crucial role in ASD, and the up-regulated inflammatory proteins may serve as potential early diagnostic biomarkers for ASD.

[Establishment and pathologic analysis of imatinib-resistant gastrointestinal stromal tumor xenografts].

OBJECTIVE: To establish and characterize imatinib-resistant gastrointestinal stromal tumor (GIST) xenografts. Further provided an ideal experimental platform through the imatinib-resistant GIST xenografts to investigate the mechanism of resistance to imatinib. METHODS: Imatinib-resistant GIST cells were injected under the skin of athymic nude mice to establish animal models of human imatinib-resistant GIST. The molecular and histopathologic features of GIST xenografts were also analysed and compared with their counterpart of cell lines. RESULTS: The xenograft tumor models had been established by subcutaneously injection of GIST cells into nude mice. Immunohistochemistry results showed CD117 expression was positive in GIST-PR2 xenograft tumor, but negative in GIST-R. In GIST-PR1, tumor areas showing rhabdomyoblastic differentiation were presented next to areas with classic GIST morphology. The rhabdomyoblastic component demonstrated consistently positivity for desmin and myogenin, whereas CD117 was completely negative. The mutation profiles of these xenograft tumors were the same as their counterpart of cell lines. CONCLUSIONS: Human GIST xenografts with mutation in c-kit have been established from imatinib-resistant GIST lines. Those models will enable further studies on mechanisms of resistance, combination therapies and allow testing of novel targeted therapies.

Transmembrane 4 L Six Family Member 1 Suppresses Hormone Receptor--Positive, HER2-Negative Breast Cancer Cell Proliferation.

Background: The prognosis of breast cancer varies according to the molecular subtype. Transmembrane 4 L six family 1 (TM4SF1) exhibits different expression patterns among the molecular subtypes of breast cancer. However, the expression profile of TM4SF1 in hormone receptor HR+HER2- breast cancer remains unclear. Methods: TM4SF1 mRNA levels were examined in major subclasses of breast cancer by analyzing The Cancer Genome Atlas (TCGA) datasets. In addition, TM4SF1 protein and mRNA levels in HR+HER2- breast cancer tissue samples were determined by immunohistochemistry and Western blot assay. The effect of TM4SF1 on cell proliferation was evaluated using MTT, colony formation, 3D organoid, and xenograft models, following the TM4SF1 overexpression or knockdown. Results: TCGA database analysis demonstrated that TM4SF1 was downregulated in breast cancer compared with the healthy adjacent breast tissue. In addition, the expression of TM4SF1 in basal-like one and the mesenchymal TNBC tissue was higher than that of the healthy adjacent breast tissue. Other types, including the luminal androgen receptor-positive TNBC tissue, expressed lower levels of TM4SF1. Immunohistochemistry and real-time quantitative PCR assays demonstrated that the TM4SF1 protein and mRNA levels were downregulated in the HR+HER2- breast cancer tissue compared with the healthy adjacent tissue. Moreover, the TM4SF1 overexpression reduced the viability of MCF-7 and ZR-75-1 breast cancer cells, whilst reducing the number of colonies and 3D-organoids formed by these cell lines. By contrast, TM4SF1 knockdown led to an increased MCF-7 cell proliferation. However, in the TNBC cell line, MDA-MB-231, TM4SF1 silencing reduced cell proliferation. In vivo, the TM4SF1 overexpression inhibited MCF-7 xenograft growth in a nude mouse model, which was associated with the downregulation of the Ki-67 expression, apoptosis induction, and inhibition of the mTOR pathway. Conclusion: TM4SF1 is downregulated in HR + HER2-breast cancer, and the overexpression of TM4SF1 suppresses cell proliferation in this cancer subtype.

Extracellular matrix metalloproteinase inducer (EMMPRIN) is present in smooth muscle cells of human aneurysmal aorta and is induced by angiotensin II in vitro.

The aim of the present study was to determine whether EMMPRIN (extracellular matrix metalloproteinase inducer) is present and is up-regulated in human aneurysmal aortas, and to assess a possible association with AngII (angiotensin II)-induced aneurysm formation. The presence of EMMPRIN was assessed in 41 surgical specimens from patients with a TAA (thoracic aortic aneurysm) (Type A aortic dissection, n=12; Type B aortic dissection, n=7; and TAA without dissection, n=7) or an AAA (abdominal aortic aneurysm, n=15) by immunohistochemistry. EMMPRIN expression in aortic aneurysm tissues was compared with 12 aortas obtained during autopsy (free of any vascular diseases), and scored for both staining intensity and the percentage of vascular cells stained. EMMPRIN protein levels in cultured human aortic SMCs (smooth muscle cells) following stimulation of AngII were analysed by Western blotting. Significant EMMPRIN immunoreactivity was detected in aortic aneurysm lesions from patients with TAAs and AAAs. In the aneurysmal wall, alpha-actin-positive SMCs were the main source of EMMPRIN. The frequency of EMMPRIN overexpression was significantly higher (P=0.026) in TAAs with dissection (68.4%) than TAAs without dissection (14.3%). AngII stimulation up-regulated the expression of EMMPIRN in cultured human aortic SMCs, which was suppressed by the addition of the AT1R (AngII type 1 receptor) antagonist losartan. In conclusion, the present study is the first to report the expression of EMMPRIN in aortic aneurysmal diseases, and we speculate that EMMPRIN may be important in the pathogenesis of these diseases. Whether these abnormalities are potential therapeutic targets deserve further investigation.

Adrenal myelolipoma within myxoid cortical adenoma associated with Conn's syndrome.

The coexistence of myelolipoma within adrenal cortical adenoma is extremely rare, for both tumors present usually as separate entities. There are only 16 such cases reported worldwide. To the best of our knowledge, the case we reported here is the first one of myxoid adrenal cortical adenoma associated with myelolipoma reported. A 32-year-old Chinese woman with 4-year history of hypertension was presented in our study. Computed tomography (CT) of the abdomen showed a large heterogeneously-enhancing mass (4.5 cm in diameter) in the left suprarenal region. Clinical history and laboratory results suggest a metabolic disorder as Conn's syndrome. The patient underwent a left adrenalectomy, and a histopathological study confirmed the mass to be a myxoid adrenal cortical adenoma containing myelolipoma. The patient was postoperatively well and discharged uneventfully. In the present case report, we also discuss the etiology of simultaneous myelolipoma and adrenal adenoma associated with Conn's syndrome, and the methods of the diagnosis and differential diagnosis.

[Interdigitating dendritic cell sarcoma/tumor: a study of 3 cases].

OBJECTIVE: To study the pathologic features, diagnosis and differential diagnosis of interdigitating dendritic cell sarcoma (IDCS). METHODS: The clinical findings, morphologic features and immunophenotype of 3 cases of IDCS were investigated. RESULTS: Gross examination showed that IDCS had a greyish-white to greyish-yellow cut surface. The site of occurrence included lung, spleen (with lymph node metastasis) and lymph node. Histologically, the tumor cells were arranged in nests, fascicles and whorls, with intimate admixture of many lymphocytes and plasma cells. They were oval to spindle in shape and contained pale eosinophilic cytoplasm, oval and sometimes grooved nuclei, small distinct nucleoli and ill-defined cell borders. Immunohistochemical study showed that the tumor cells expressed S-100 protein. CONCLUSIONS: IDCS is a rare type of histiocytic and dendritic cell malignancy with distinctive morphologic findings. It needs to be distinguished from follicular dendritic cell sarcoma, inflammatory pseudotumor, Langerhans' cell histiocytosis, malignant melanoma, undifferentiated carcinoma and anaplastic large cell lymphoma. Immunohistochemical staining for S-100 protein is helpful in confirming the diagnosis.

GRP137 promotes cell proliferation and metastasis through regulation of the PI3K/AKT pathway in human ovarian cancer.

PURPOSE: Ovarian cancer is one of the leading causes of death for women worldwide. The present study aims to investigate the role of G protein-coupled receptor 137 (GPR137) in the biological activities of ovarian cancer cells. METHODS: (QUERY: Please supply Methods for Abstract) RESULTS: G protein-coupled receptor 137 was highly expressed in clinical ovarian cancer tissues and exhibited the highest protein levels in SKOV3 cells and OVCAR3 cells. Knockdown of GPR137 caused significant decreases in cell proliferative rates and colony formation abilities in SKOV3 cells and OVCAR3 cells and also inhibited the in vivo tumorigenesis in a xenograft model. It was observed that knockdown of GPR137 inhibited cell motility by up to 40% in SKOV3 cells and approximately 65% in OVCAR3 cells in wound-healing assay. Cell migration abilities were consistently inhibited by 68.2% in SKOV3 cells and 59.3% in OVCAR3 cells, whereas cell invasion abilities were inhibited by 64.0% and 74.2% in SKOV3 and OVCAR3 cells, respectively, after knockdown of GPR137. When GPR137 was depleted, epithelial markers were increased, while mesenchymal markers decreased. CONCLUSIONS: Our data suggest that GPR137 plays pro-oncogenic roles in ovarian cancer via regulation of the PI3K/AKT pathway. These observations might pave new insights into therapeutic strategies against human ovarian cancer.

Aloesin Suppresses Cell Growth and Metastasis in Ovarian Cancer SKOV3 Cells through the Inhibition of the MAPK Signaling Pathway.

Aloesin is an active constituent of the herb aloe vera and plays a crucial role in anti-inflammatory activity, ultraviolet protection, and antibacterium. We investigated the role and possible mechanisms of aloesin in the cell growth and metastasis of ovarian cancer. It was found that aloesin inhibited cell viability and cell clonality in a dose-dependent manner. It arrests the cell cycle at the S-phase and induced apoptosis in SKOV3 cells. In an in vivo experiment, it was observed that aloesin inhibited tumor growth. Moreover, it inhibited migration and invasion of cancer in SKOV3 cells. Interestingly, members from the mitogen-activated protein kinase (MAPK) signaling family became less phosphorylated as the aloesin dose increased. This suggests that aloesin exerts its anticancer effect through the MAPK signaling pathway. Our data also highlights the possibility of using aloesin as a novel therapeutic drug for ovarian cancer treatment.

Long Noncoding RNA MIR4697HG Promotes Cell Growth and Metastasis in Human Ovarian Cancer.

Ovarian cancer is one of the three most common gynecological malignant tumors worldwide. The prognosis of patients suffering from this malignancy remains poor because of limited therapeutic strategies. Herein, we investigated the role of a long noncoding RNA named MIR4697 host gene (MIR4697HG) in the cell growth and metastasis of ovarian cancer. Results showed that the transcriptional level of MIR4697HG in cancerous tissues increased twofold compared with that in adjacent noncancerous tissues. MIR4697HG was differentially expressed in ovarian cancer cell lines, with the highest levels in OVCAR3 and SKOV3 cells. MIR4697HG knockdown by specific shRNA significantly inhibited cell proliferation and colony formation in both OVCAR3 and SKOC3 cells. Consistently, in a xenograft model of ovarian cancer, MIR4697HG depletion also significantly restricted tumor volumes and weights. Furthermore, MIR4697HG knockdown inhibited cell migration and invasion capacities. Invasion ability was inhibited by 58% in SKOV3 cells and 40% in OVCAR3 cells, and migration ability was inhibited by 73% in SKOV3 cells and 62% in OVCAR3 cells after MIR4697HG knockdown. MIR4697HG knockdown also caused a decrease in matrix metalloprotease-9, phosphorylated ERK, and phosphorylated AKT. These data suggested that MIR4697HG promoted ovarian cancer growth and metastasis. The aggressive role of MIR4697HG in ovarian cancer may be related to the ERK and AKT signaling pathways.

Expression and prognostic role of MEKK3 and pERK in patients with renal clear cell carcinoma.

Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is an important serine/threonine protein kinase and a member of the MAPK family. MEKK3 can effectively activate the MEK/ERK signaling pathway and promote an autocrine growth loop critical for tumor genesis, cell proliferation, terminal differentiation, apoptosis and survival. To explore the relationship between MEKK3 and cell apoptosis, clinicopathology and prognosis, we characterize the expression of MEKK3, pERK and FoxP3 in the renal clear cell carcinoma (RCCC). Protein expression was detected by tissue microarray and immunochemistry in 46 cases of RCCC and 28 control cases. Expression levels of CD3+ ,CD3+CD4+,CD3+CD8+,CD4+CD25+, CD4+CD25+ FoxP3+ were assessed by flow cytometry and analyzed for their association with pathological factors, correlation and prognosis in RCCC. Expression of MEKK3, pERK and FoxP3 was significantly up-regulated in RCCC as compared to control levels (p<0.01), associated with pathological grade (p<0.05)and clinical stage (p<0.05). CD4+CD25+ Foxp3+ Treg cells were also significantly increased in RCCC patients (p<0.05). Cox multivariate regression analysis showed that MEKK3, pERK expression and patholigical stage were independent prognostic factors in patients with RCCC (p<0.05). MEKK3 can be used as an important marker of early diagnosis and prognostic evaluation in RCCC. It may be associated with imbalance of anti-tumor immunity and overexpression of pERK. Expression of MEKK3 and pERK are significantly increased in RCCC, with protein expression and clinical stage acting as independent prognostic factors.

Schwann cell apoptosis in Wallerian-degenerated sciatic nerve of the rat.

OBJECTIVE: To investigate systematically Schwann cell apoptosis in Wallerian-degenerated sciatic nerve of the rat, and evaluate its time-related feature. METHODS: Ninety-five SD rats were divided randomly into one normal group (8 rats) and 11 experimental groups (66 rats, 6 in each). Both hind legs of each rat in experimental groups were randomly divided into test leg (sciatic nerve transected) and control one (nerve uninjured). All test legs constituted a test group and all control legs constituted a control one. After operation, all rats were respectively sacrificed at 1 h, 6 h, 12 h, 24 h, 2 d, 3 d, 4 d, 8 d, 14 d, 21 d, and 30 d. We analyzed the specimens of mid-distal sciatic nerve, especially the morphological changes of the nerve, the different expression levels of S-100 protein and apoptosis-related proteins such as Bcl-2, Bax, and Fas in Schwann cells. The TUNEL method was used to detect the apoptotic rate of Schwann cells. RESULTS: (1) The test group showed Wallerian degeneration. The number of Schwann cells began to decrease at 24 h, obviously decreased on day 3 and 4, then began to increase from day 8 and formed Bungner belt after 14 days. (2) Schwann cells generally expressed S-100 at a low level in all groups. The control group was not significantly different from the normal group. The test group had statistical significance at 1 h and day 21. (3) As an inhibitory gene protein of Schwann cell apoptosis, Bcl-2 positive rates in the control and test groups apparently elevated and were statistically different from the normal group. (4) As a promotive gene protein of Schwann cell apoptosis, the control and test groups expressed Bax at a high level and were statistically different from the normal group. (5) As a promotive gene protein of Schwann cell apoptosis, Fas positive rate in control group was slightly elevated, but had no statistical significance compared with the normal group. Fas positive rate in test group continuously elevated in a fluctuant way, with highly statistical significance compared with the normal group. (6) TUNEL detection further proved that Schwann cell apoptosis rarely existed in the normal group, and the left sciatic nerve had no statistical significance compared with the right sciatic nerve. While the test group showed lots of apoptotic nuclei at 6 h, 2 d, 4 d, and 21 d. It had highly statistical significance compared with the normal group. CONCLUSIONS: Schwann cell apoptosis does exist in Wallerian-degenerated sciatic nerve of the rat after transection. Schwann cell apoptosis and its apoptotic genes expression have a time-related feature.

MEKK3 and survivin expression in cervical cancer: association with clinicopathological factors and prognosis.

Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3) is an important protein kinase and a member of the MAPK family, which regulates cellular responses to environmental stress and serves as key integration points along the signal transduction cascade that not only link diverse extracellular stimuli to subsequent signaling molecules but also amplify the initiating signals to ultimately activate effector molecules and induce cell proliferation, differentiation and survival. To explore the relationship between MEKK3 and cell apoptosis, clinicopathology and prognosis, we characterize the expression of MEKK3 and survivin in cervical cancer. MEKK3 and survivin expression was measured by RT-PCR and Western blotting of fresh surgical resections from 30 cases of cervical cancer and 25 cases of chronic cervicitis. Protein expression was detected by tissue microarray and immunochemistry (En Vision) in 107 cases of cervical cancer, 86 cases of cervical intraepithelial neoplasia (CIN), and 35 cases of chronic cervicitis. Expression patterns were analyzed for their association with clinicopathological factors and prognosis in cervical cancer. Expression of MEKK3 and survivin mRNA was significantly higher in cervical cancer than in the controls (p<0.05). MEKK3 and survivin expression differed significantly between cervical carcinoma, CIN, and cervicitis (p<0.05) and correlated with clinical stage, infiltration depth, and lymph node metastasis (p<0.05). MEKK3 expression was positively correlated with survivin (p<0.05). Kaplan-Meier survival analysis showed that MEKK3 and survivin expression, lymph node metastasis, depth of invasion, and FIGO stage reduce cumulative survival. Cox multivariate regression analysis showed that MEKK3, survivin, and clinical staging are independent prognostic factors in cervical cancer (p<0.05). Expression of MEKK3 and survivin are significantly increased in cervical cancer, their overexpression participating in the occurrence and development of cervical cancer, with protein expression and clinical staging acting as independent prognostic factors for patients with cervical cancer.

Expression and function analysis of mitotic checkpoint genes identifies TTK as a potential therapeutic target for human hepatocellular carcinoma.

The mitotic spindle checkpoint (SAC) genes have been considered targets of anticancer therapies. Here, we sought to identify the attractive mitotic spindle checkpoint genes appropriate for human hepatocellular carcinoma (HCC) therapies. Through expression profile analysis of 137 selected mitotic spindle checkpoint genes in the publicly available microarray datasets, we showed that 13 genes were dramatically up-regulated in HCC tissues compared to normal livers and adjacent non-tumor tissues. A role of the 13 genes in proliferation was evaluated by knocking them down via small interfering RNA (siRNA) in HCC cells. As a result, several mitotic spindle checkpoint genes were required for maintaining the proliferation of HCC cells, demonstrated by cell viability assay and soft agar colony formation assay. Then we established sorafenib-resistant sublines of HCC cell lines Huh7 and HepG2. Intriguingly, increased TTK expression was significantly associated with acquired sorafenib-resistance in Huh7, HepG2 cells. More importantly, TTK was observably up-regulated in 46 (86.8%) of 53 HCC specimens. A series of in vitro and in vivo functional experiment assays showed that TTK overexpression promoted cell proliferation, anchor-dependent colony formation and resistance to sorafenib of HCC cells; TTK knockdown restrained cell growth, soft agar colony formation and resistance to sorafenib of HCC cells. Collectively, TTK plays an important role in proliferation and sorafenib resistance and could act as a potential therapeutic target for human hepatocellular carcinoma.