lncRNA ZNF674-AS1 inhibits the migration, invasion and epithelial-mesenchymal transition of thyroid cancer cells by modulating the miR-181a/SOCS4 axis.

Thyroid cancer (TC) is a very common endocrine cancer worldwide. Further understanding and revealing the molecular mechanism underlying thyroid cancer are indispensable for the development of effective diagnosis and treatments. Long non-coding RNAs (lncRNAs), a series of non-coding RNAs with a length of >200 nts, have been regarded as crucial regulators of many cancers playing a tumor suppressive or oncogenic role, depending on circumstances. lncRNA ZNF674-AS1 was reported to be abnormally expressed in TC, but the exact mechanism remains unclear. This study aims to probe the mechanism and roles of ZNF674-AS1 in TC. The expression patterns of RNAs and proteins were determined via qRT-PCR and western blotting, respectively. Cell proliferation, migration and invasion were detected using MTT and Transwell assays. ZNF674-AS1 and SOCS4 expression were remarkably reduced while miR-181a was upregulated in TC tissues and cells. Enforced expression of ZNF674-AS1 inhibited proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) in vitro and reduced tumour growth in vivo. Mechanistic assays verified that ZNF674-AS1 directly interacted with miR-181a to increase SOCS4 expression. In addition, miR-181a overexpression aggravated proliferation, metastasis and EMT by inhibiting SOCS4. Interestingly, inhibition of miR-181a diminished the promoting effects of ZNF674-AS1 silencing on the malignant behaviours of TC cells. These data illustrated that ZNF674-AS1 alleviated TC progression by modulating the miR-181a/SOCS4 axis (graphical abstract), further suggesting that ZNF674-AS1 might be used as a therapheutic target in TC treatment.

miR-195-5p regulates cell proliferation, apoptosis, and invasion of thyroid cancer by targeting telomerase reverse transcriptase.

In most human primary cancers, the expression, or telomerase activity, of telomerase reverse transcriptase (TERT) is detectable. However, the mechanism ofTERTactivity within oncogenesis of thyroid cancer remains largely unknown. In this study, we identified miR-195-5p as having involvement in cell proliferation, apoptosis, and invasion in human thyroid cancer. MTT was used to measure cell proliferation, Transwell chamber was used to measure invasion. Western blotting was used to detect the expressions of TERT, PCNA, and Ki67. Target gene prediction software predicted that TERT may be the target gene of miR-195-5p. Luciferase reporting system was used to identify the targeting relationship. A significant increase of in TERT expression was observed by immunohistochemistry compared with normal tissue, however, a decrease in miR-195-5p expression using qRT-PCRand western blot compared with normal cells. Functional analysis demonstrates that miR-195-5p negatively correlated withTERTand inhibitedTERTexpression through its interaction with theTERT3'-untranslatedregion (3'-UTR). Overexpression of miR-195-5p was shown to inhibit proliferation and invasion, and promote apoptosis of CAL-62 thyroid cancer cells. miR-195-5p-mediatedeffects were rescued by the overexpression ofTERT. Altogether, our data demonstrate that miR-195-5p regulates cell proliferation, apoptosis, and invasion in human thyroid cancer viaTERT, providing evidence of a new potential therapeutic target for further investigation.

Integrated tissue proteome and metabolome reveal key elements and regulatory pathways in cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (CSCC) is a widespread malignancy but has a very low long-term survival rate for patients at the metastatic stage. Therefore, it is urgent to identify prognostic biomarkers for CSCC and improve our understanding of disease progression. Here we took advantage of a data-independent acquisition (DIA)-based nano liquid chromatography equipped with an orbitrap mass spectrometry (nLC-MS/MS) and ultraperformance LC coupled to a time-of-flight tandem MS (UPLC-TOF-MS/MS) technique to analyze cancer and corresponding noncancerous tissues from 20 CSCC patients for integrated proteomic and metabolomic analyses. Overall, 6241 tissue proteins were detected, while 136 proteins were significantly expressed in CSCC tissues. Further functional analysis revealed that various biological processes were highly enriched and participated in the pathogenesis of CSCC, especially DNA damage responses. Moreover, 641 named metabolites in total were identified, among which 181 were significantly changed in CSCC tissues. A total of 101 pathways were significantly enriched including apoptosis, autophagy, PI3K-Akt and mTOR signaling pathways. Interestingly, two pathways, protein digestion & absorption and platelet activation were both enriched in proteomic and metabolomic studies involving 5 proteins and 11 metabolites. Accordingly, a four-metabolite panel consisting of arachidonate, glutamine, glutamic acid, and proline (all area under the curve (AUC) values more than 0.9) was developed with a high accuracy (0.971) to distinguish the 20 detected cancer tissues from their noncancerous tissues. Collectively, our work highlighted the key elements and regulatory pathways involved in the pathogenesis of CSCC. More importantly, the present study not only provided potential biomarkers for the early diagnosis of CSCC, but also expanded our knowledge of the physiopathology of the disease. SIGNIFICANCE: CSCC is the second most common human cancer but has few treatment options and few sensitive biomarkers for diagnosis. Here we comprehensively revealed its molecular characteristics by performing integrated tissue proteomic and metabolomic analyses. Significantly distinct profiles and certain enriched pathways including DNA damage responses were identified as associated with CSCC. Moreover, protein digestion & absorption and platelet activation were both enriched in the proteome and metabolome. These identified molecular changes probably play significant roles in CSCC development. Finally, we developed a four-metabolite panel to distinguish CSCC with high accuracy. Overall, our data not only provided potential diagnostic biomarkers, but also extended knowledge on the pathogenesis of CSCC.

LncRNA NCK1-AS1 in plasma distinguishes oral ulcer from early-stage oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) at early stages can be misdiagnosed as an oral ulcer (OU) due to similar symptoms, such as chronic and indurated ulcer. LncRNA NCK1-AS1 has been characterized as a key player in cervical cancer, while its role in OSCC is unknown. METHODS: All participants were selected at Jiangxi Province Tumor Hospital from December 2016 to December 2018. Expression levels of NCK1-AS1 and miR-100 in plasma from both OSCC and OU patients were measured by RT-qPCR. Diagnostic analysis was performed through ROC curve. Potential interactions between NCK1-AS1 and miR-100 were detected by cell transfection experiments. Cell invasion and migration were assessed by Transwell assays. RESULTS: The expression of NCK1-AS1 was upregulated in early-stage OSCC patients but not in OU patients. Upregulation of NCK1-AS1 distinguished OSCC patients from OU patients. The expression of miR-100 was inversely correlated with the expression of NCK1-AS1. Overexpression of NCK1-AS1 was followed by promoted OSCC cell invasion and migration. Overexpression of miR-100 did not affect the expression of NCK1-AS1 but inhibited the role of NCK1-AS1. CONCLUSIONS: Therefore, NCK1-AS1 may promote the metastasis of OSCC by downregulating miR-100.

Vibrio parahaemolyticus infection impaired intestinal barrier function and nutrient absorption in Litopenaeus vannamei.

The intestine is the primary target of pathogenic microbes during invasion. However, the interaction of Vibrio parahaemolyticus (V. parahaemolyticus) with intestinal epithelial cells and its effects on the intestinal function of Litopenaeus vannamei (L. vannamei) are poorly studied. Therefore, the aim of this study was to investigate the influence of V. parahaemolyticus infection on intestinal barrier function and nutrient absorption in L. vannamei. In the present study, a total of 90 shrimp were randomly divided into two groups including the control group and V. parahaemolyticus infection group (final concentration of 1 × 105 CFU/mL), with three replicates per group. The result showed that compared with the control group, V. parahaemolyticus infection increased (P < 0.05) serum diamine oxidase activity and endotoxin quantification, and down-regulated (P < 0.05) the mRNA levels of intestinal peroxinectin, integrin, midline fasciclin at 48 h and 72 h; V. parahaemolyticus infection decreased (P < 0.05) the mRNA expression of intestinal amino acid transporter (CAT1, EAAT3 and ASCT1) and glucose transporter (SGLT-1, GLUT) at 24 h, 48 h and 72 h, and increased (P < 0.05) serum glucose and amino acid (Asp, Thr, Ser, Glu, Gly, Ala, Val, Ile, Leu, Tyr, Phe, Lys, His and Arg) concentration at 24 h. The results indicated that V. parahaemolyticus infection increased intestinal permeability, inhibited absorption of glucose and amino acid in L. vannamei.

LncRNA WT1-AS Downregulates Survivin by Upregulating miR-203 in Papillary Thyroid Carcinoma.

OBJECTIVE: This study aimed to assessment the functions of lncRNA WT1-AS in papillary thyroid carcinoma (PTC). METHODS: Expression levels of WT1-AS in PTC and non-tumor tissues from 66 PTC patients were measured and compared by performing qPCR and paired t test, respectively. Cell proliferation (CCK-8) assay was performed to evaluate the effects of the overexpression of WT1-AS, miR-203 and survivin on the proliferation of IHH-4 (a human PTC cell line) cells. RESULTS: We found that WT1-AS was significantly downregulated in PTC and associated with clinical stages. In PTC tissues, WT1-AS was negatively correlated with survivin but positively correlated with miR-203. In PTC cells, WT1-AS overexpression led to significantly upregulated miR-203 and downregulated survivin. MiR-203 overexpression failed to affect WT1-AS but downregulated survivin. Cell proliferation assay showed that overexpression of WT1-AS and miR-203 led to decreased, while survivin overexpression led to increased proliferation of PTC cells. In addition, survivin overexpression attenuated the effects of WT1-AS and miR-203 overexpression. CONCLUSION: Therefore, WT1-AS may downregulate survivin by upregulating miR-203 in PTC to inhibit cancer cell proliferation.

The levels of NF-κB p50 and NF-κB p65 play a role in thyroid carcinoma malignancy in vivo.

Background To investigate the relationship between the levels of nuclear factor (NF)-κB p50 and NF-κB p65 and tumour characteristics in patients with thyroid carcinoma. Methods This prospective study enrolled consecutive patients with thyroid carcinoma. Tumour samples were collected and the levels of NF-κB p50 and NF-κB p65 protein and mRNA were measured using immunohistochemistry and quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Results A total of 73 patients with thyroid carcinoma were included in the study (20 males; 53 females; mean ± SD age, 44.8 ± 12.7 years, range, 18-76 years). There were no significant differences in sex, age and pathological type between the NF-κB p50 positive group and the NF-κB p50 negative group, but tumour diameter and lymph node metastasis were significantly higher in the NF-κB p50 positive group compared with the NF-κB p50 negative group. Similar findings were observed for NF-κB p65. The levels of NF-κB p50 were positively correlated with NF-κB p65 in samples of thyroid carcinoma ( rs = 0.653). Conclusion The levels of NF-κB p50 and NF-κB p65 in samples of thyroid carcinoma were positively associated with tumour diameter and the presence of lymph node metastasis.

Anemonin improves intestinal barrier restoration and influences TGF-ß1 and EGFR signaling pathways in LPS-challenged piglets.

The present study was aimed at investigating whether dietary anemonin could alleviate LPS-induced intestinal injury and improve intestinal barrier restoration in a piglet model. Eighteen 35-d-old pigs were randomly assigned to three treatment groups (control, LPS and LPS+anemonin). The control and LPS groups were fed a basal diet, and the LPS + anemonin group received the basal diet + 100 mg anemonin/kg diet. After 21 d of feeding, the LPS- and anemonin-treated piglets received i.p. administration of LPS; the control group received saline. At 4 h post-injection, jejunum samples were collected. The results showed that supplemental anemonin increased villus height and transepithelial electrical resistance, and decreased crypt depth and paracellular flux of dextran (4 kDa) compared with the LPS group. Moreover, anemonin increased tight junction claudin-1, occludin and ZO-1 expression in the jejunal mucosa, compared with LPS group. Anemonin also decreased TNF-α, IL-6, IL-8 and IL-1ß mRNA expression. Supplementation with anemonin also increased TGF-ß1 mRNA and protein expression, Smad4 and Smad7 mRNA expressions, and epidermal growth factor and epidermal growth factor receptor (EGFR) mRNA expression in the jejunal mucosa. These findings suggest that dietary anemonin attenuates LPS-induced intestinal injury by improving mucosa restoration, alleviating intestinal inflammation and influencing TGF-ß1 canonical Smads and EGFR signaling pathways.

L-cysteine protects intestinal integrity, attenuates intestinal inflammation and oxidant stress, and modulates NF-κB and Nrf2 pathways in weaned piglets after LPS challenge.

In this study we investigated whetherL-cysteine (L-cys) could alleviate LPS-induced intestinal disruption and its underlying mechanism. Piglets fed with anL-cys-supplemented diet had higher average daily gain.L-cys alleviated LPS-induced structural and functional disruption of intestine in weanling piglets, as demonstrated by higher villus height, villus height (VH) to crypt depth (CD) ratio, and transepithelial electrical resistance (TER) and lower FITC-dextran 4 (FD4) kDa flux in jejunum and ileum. Supplementation withL-cys up-regulated occludin and claudin-1 expression, reduced caspase-3 activity and enhanced proliferating cell nuclear antigen expression of jejunum and ileum relative to LPS group. Additionally,L-cys suppressed the LPS-induced intestinal inflammation and oxidative stress, as demonstrated by down-regulated TNF-α, IL-6 and IL-8 mRNA levels, increased catalase, superoxide dismutase, glutathione peroxidase activity, glutathione (GSH) contents and the ratio of GSH and oxidized glutathione in jejunum and ileum. Finally, a diet supplemented withL-cys inhibited NF-κB(p65) nuclear translocation and elevated NF erythroid 2-related factor 2 (Nrf2) translocation compared with the LPS group. Collectively, our results indicated the protective function ofL-cys on intestinal mucosa barrier may closely associated with its anti-inflammation, antioxidant and regulating effect on the NF-κB and Nrf2 signaling pathways.

Zinc oxide influences intestinal integrity, the expressions of genes associated with inflammation and TLR4-myeloid differentiation factor 88 signaling pathways in weanling pigs.

This study explored whether zinc oxide (ZnO) supplementation could alleviate weanling-induced intestinal injury through TLR and NOD-like receptor signaling pathways. Twelve early-weanling piglets were allotted to two dietary treatments (control vs 2200 mg Zn/kg from ZnO) for 1 wk. The results showed that supplemental ZnO improved daily gain and feed intake, decreased post weaning scour scores, increased villus height and villus height:crypt depth ratio at the jejunal mucosa, and decreased diamine oxidase activity and endotoxin concentration in plasma. The intestinal mRNA levels of TLR4 and its downstream signals, including MyD88, IL-1 receptor-associated kinase 1 and TNF-α receptor-associated factor 6, were decreased, and the expressions of intestinal pro-inflammatory cytokines and chemokines were decreased simultaneously in the ZnO-supplemented piglets. Although NF-κB p65 mRNA abundance was not affected by ZnO supplementation, NF-κB p65 protein expression was down-regulated by ZnO. However, ZnO supplementation had no effect on intestinal expressions of NOD1 and NOD2, and their adaptor molecule receptor-interacting serine/threonine-protein kinase 2, as well as protein expressions of caspase-3 and heat shock protein 70. The results indicated that the protective effects of ZnO on intestinal integrity were closely related to decreasing the expressions of genes associated with inflammation through inhibiting the TLR4-MyD88 signaling pathways.

[Change of matrix metalloproteinase-1 and matrix metalloproteinase-7 in serum and bronchoalveolar lavage fluid of patients with idiopathic pulmonary fibrosis and sarcoidosis].

OBJECTIVE: To detect the levels of matrix metalloprotease (MMP)-1 and MMP-7 in the serum and the bronchoalveolar lavage fluid (BALF) of patients with idiopathic pulmonary fibrosis (IPF) and sarcoidosis (Stage II), and therefore to investigate the significance of these changes in the pathogenesis of IPF. METHODS: Forty-four clinically confirmed cases of IPF were recruited, with the patients' age ranging from 46 to 70 years (58 ± 9 years). Twenty patients with sarcoidosis, aged 35 to 65 (50 ± 12) years, were also studied. Enzyme-linked immunoabsorbent assay was used to detect the levels of MMP-1 and MMP-7 in the serum and the BALF samples. RESULTS: In the serum of patients with IPF, the level of MMP-1 [3.78 (0.14 - 13.44) µg/L] was lower than that in patients with sarcoidosis [7.79 (4.67 - 10.68) µg/L (z = -3.53, P < 0.01)], but the level of MMP-7 [7.83 (3.57 - 14.37) µg/L] was higher than that in patients with sarcoidosis [4.04 (0.06 - 9.94) µg/L (z = -3.84, P < 0.01)]. In the BALF of patients with IPF, the level of MMP-1 [1.09 (0.04 - 5.14) µg/L] was lower than that in patients with sarcoidosis [2.08 (0.05 - 4.16) µg/L (z = -1.53, P > 0.05)], but the level of MMP-7 [3.75 (1.10 - 9.87)µg/L] was higher than that in patients with sarcoidosis [1.16 (0.02 - 4.47) µg/L (z = -5.33, P < 0.01)]. The serum level of MMP-7 in patients with IPF was negatively correlated with the diffusing capacity of carbon monoxide(r = -0.56, P < 0.01) and the percentage of neutrophils (r = -0.47, P < 0.01). The level of MMP-7 in the BALF showed a negative correlation with diffusing capacity of carbon monoxide (r = -0.31, P < 0.05). CONCLUSIONS: The results suggest that MMP-1 may be increased in the inflammatory phase as compared to the matrix remodeling phase of lung fibrosis, while MMP-7 may be increased in the matrix remodeling phase rather than in the inflammatory phase. MMP-7 may act as an important indicator for the severity of IPF.

Hematopoiesis of immature myeloid dendritic cells in stroma-dependent spleen long-term cultures occurs independently of NF-KB/RelB function.

OBJECTIVE: The nuclear factor-kappaB (NF-KB)/RelB transcription factor plays an essential role in development of some dendritic cell (DC) subsets in mice. In this laboratory, immature myeloid DC are produced in vitro in a stroma-dependent murine spleen long-term culture (LTC) system. In LTC, DC differentiate from hematopoietic progenitors maintained within the stromal cell matrix. Expression and function of RelB in development of LTC-DC has been investigated, with a view to assessing the relationship between DC produced in this system and other known subsets of DC. MATERIALS AND METHODS: RelB expression by LTC-DC was confirmed by detection of protein by Western blotting, RNA by reverse transcription polymerase chain reaction, and nuclear protein with DNA-binding function in electrophoretic mobility shift assays. The role of RelB in cell development was assessed by addition of antisense RelB oligonucleotides into LTC and colony assays established above STX3 stromal cells. RelB(-/-) mice were also examined for ability to produce LTC, and for presence of DC progenitors in spleen and bone marrow that can generate DC when overlaid on STX3 in cocultures. RESULTS: Functional RelB was detected in both LTC-DC and in STX3 stromal cells. A critical role for RelB in DC differentiation from spleen progenitors was confirmed, because antisense RelB oligonucleotides specifically and completely inhibited production of large differentiated myeloid DC in LTC. Further investigation using RelB(-/-) mice revealed that RelB expression by stromal cells rather than hematopoietic cells was required for production of LTC-DC. This was evidenced by a combination of factors, including 1) inability to generate productive LTC from RelB(-/-) mice; 2) presence of DC precursors in RelB(-/-) bone marrow and spleen, which could produce DC in stromal cocultures; and 3) increased myeloid precursor frequency among RelB(-/-) spleen cells over RelB(+/+) control cell populations. CONCLUSION: Specific development of fully differentiated, but immature myeloid CD11c(+)CD11b(+)MHC-CII(-)CD8alpha(-)CD40(-) DC in spleen LTC is dependent on expression of activated NF-kappaB/Rel-B. However, this appears to relate to stromal cell function rather than to the function of hematopoietic cells. Altogether these data confirm the importance of splenic stromal cells in myelopoiesis leading to development of immature DC as produced in LTC.

Neuroendoscopic management of symptomatic septum pellucidum cysts.

OBJECTIVE: Ten rare cases of symptomatic septum pellucidum cysts in patients who underwent endoscopic fenestration are described. The approaches and techniques used in the management of these cysts and the endoscopic surgical indications are discussed. CLINICAL PRESENTATION: In the past 5 years, 10 patients (age range, 3-60 yr) with symptomatic septum pellucidum cysts underwent neuroendoscopic fenestration. The most common symptom was intermittent headache (seven patients) accompanied by dizziness, vomiting, and epileptic seizures. Two patients presented with epileptic seizures. One patient presented with abnormally increased head circumference. Magnetic resonance imaging scans of 10 patients showed septum pellucidum cysts, two with hydrocephalus, and two with pituitary microadenoma. INTERVENTION: All 10 patients underwent endoscopic fenestration with a rigid endoscope via a frontal approach. Eight cases were performed freehand. Two cases were assisted by a frameless neuronavigation system. Postoperatively, the mass effect of the cysts and the symptoms resolved immediately, and computed tomographic or magnetic resonance imaging scans showed significant decrease in the cyst size and no recurrence during follow-up. Ventricular sizes in the two patients with hydrocephalus were normal. CONCLUSION: Neuroendoscopic pellucidotomy could be an effective, safe, and convenient therapeutic method for symptomatic septum pellucidum cysts. This approach might provide communication between the cyst and the ventricular system, thus avoiding shunting or craniotomy. We consider that it is appropriate to use the rigid endoscope via the frontal approach. It is helpful to fill the ventricles with lactated Ringer's solution and leave an external drain after surgery.