Immune cell signatures and inflammatory mediators: unraveling their genetic impact on chronic kidney disease through Mendelian randomization.

Prior research has established associations between immune cells, inflammatory proteins, and chronic kidney disease (CKD). Our Mendelian randomization study aims to elucidate the genetic causal relationships among these factors and CKD. We applied Mendelian randomization using genetic variants associated with CKD from a large genome-wide association study (GWAS) and inflammatory markers from a comprehensive GWAS summary. The causal links between exposures (immune cell subtypes and inflammatory proteins) and CKD were primarily analyzed using the inverse variance-weighted, supplemented by sensitivity analyses, including MR-Egger, weighted median, weighted mode, and MR-PRESSO. Our analysis identified both absolute and relative counts of CD28 + CD45RA + CD8 + T cell (OR = 1.01; 95% CI = 1.01-1.02; p < 0.001, FDR = 0.018) (OR = 1.01; 95% CI = 1.00-1.01; p < 0.001, FDR = 0.002), CD28 on CD39 + CD8 + T cell(OR = 0.97; 95% CI = 0.96-0.99; p < 0.001, FDR = 0.006), CD16 on CD14-CD16 + monocyte (OR = 1.02; 95% CI = 1.01-1.03; p < 0.001, FDR = 0.004) and cytokines, such as IL-17A(OR = 1.11, 95% CI = 1.06-1.16, p < 0.001, FDR = 0.001), and LIF-R(OR = 1.06, 95% CI = 1.02-1.10, p = 0.005, FDR = 0.043) that are genetically predisposed to influence the risk of CKD. Moreover, the study discovered that CKD itself may causatively lead to alterations in certain proteins, including CST5(OR = 1.16, 95% CI = 1.09-1.24, p < 0.001, FDR = 0.001). No evidence of reverse causality was found for any single biomarker and CKD. This comprehensive MR investigation supports a genetic causal nexus between certain immune cell subtypes, inflammatory proteins, and CKD. These findings enhance the understanding of CKD's immunological underpinnings and open avenues for targeted treatments.

Targeting PEAK1 sensitizes anaplastic thyroid carcinoma cells harboring BRAFV600E to Vemurafenib by Bim upregulation.

Pseudopodium-enriched atypical kinase 1 (PEAK1) has been demonstrated to be upregulated in human malignancies and cells. Enhanced PEAK1 expression facilitates tumor cell survival and chemoresistance. However, the role of PEAK1 inhibition to anaplastic thyroid carcinoma cell (ATC) and vemurafenib resistance is still unknown. Here, we observed that targeting PEAK1 inhibited cell viability and colony formation, but not cell apoptosis in both of the 8505C and Hth74 cells in vitro. Targeting PEAK1 sensitized 8505C and Hth74 cells to vemurafenib by inducing cell apoptosis, and thereby decreasing cell viability. Mechanistically, vemurafenib treatment upregulated PEAK1 expression. Combined PEAK1 depletion and Vemurafenib treatment upregulated Bim expression. Targeting PEAK1 sensitized vemurafenib-induced apoptosis by upregulating Bim. In conclusion, vemurafenib resistance in ATC cells harboring BRAFV600E is associated with PEAK1 activation, resulting in the inhibition of pro-apoptotic Bim protein. Therefore, targeting PEAK1 may be an effective strategy to sensitize ATC harboring BRAFV600E to vemurafenib.

Association of Serum Potassium Variability With 60-day Mortality and Cardiovascular Events after COVID-19 Infection in Maintenance Hemodialysis Patients.

Objective: This study aimed to investigate the association between serum potassium variability and 60-day mortality and cardiovascular disease (CVD) in maintenance hemodialysis (MHD) patients following the coronavirus disease 2019 (COVID-19) infection. Methods: We conducted a retrospective study on MHD patients treated at the affiliated hospital of Qingdao University hemodialysis center who were infected with the novel coronavirus between December 1, 2022, and January 31, 2023. Baseline characteristics of patients were collected from electronic medical records. Kaplan-Meier survival analysis was used to obtain patient survival probabilities, and multivariate Cox hazard regression models and binary Logistic regression models were used to obtain hazard ratios (HR), odds ratios (OR), and 95% confidence intervals (95% CI) between exposure and outcomes. Results: A total of 296 patients were included in this study, with a mean age of 57.2±16.3 years, and 59.8% were male. The 60-day mortality rate was 10.8%, and the incidence of CVD was 32.8%. Kaplan-Meier curves showed that a higher potassium variability coefficient was associated with higher all-cause mortality (P = 0.024). After adjusting for potential confounders, multivariate Cox regression analysis showed that the HR for 60-day mortality in the Q4 group compared to the Q1 group was 2.06 (95% CI = 1.03-4.09, P = 0.040), and binary Logistic regression analysis showed that the OR for 60-day CVD in the Q4 group compared to the Q1 group was 4.09 (95% CI = 1.52-10.97, P = 0.005). Conclusion: Increased serum potassium variability in MHD patients after COVID-19 infection significantly increased the likelihood of 60-day mortality and CVD.

Expression and Clinical Significance of Th1/Th2/Th17 Cytokines and Lymphocyte Subsets in PCNSL.

Purpose: Primary central nervous system lymphoma (PCNSL) responds favorably to radiation, chemotherapy and targeted drug therapy. However survival is usually worse, the treatment-related drug resistance and recurrence are still clinical problems to be solved urgently. Studies have shown that cytokines are expressed in varying degrees in patients with lymphoma, which is significantly related to the progression, poor prognosis and drug resistance of lymphoma. We explore the expression and clinical significance of Th1/Th2/Th17 cytokines and lymphocyte subsets in patients with PCNSL to provide a more sufficient theoretical basis for its diagnosis and treatment. Patients and Methods: We measured and analysed the levels of Th1/Th2/Th17 cytokines and the distribution of lymphocyte subsets (including Treg cells, CD3+, CD4+, CD8+, CD19+, and CD4+/CD8+) in 39 patients with PCNSL and 96 patients with diffuse large B-cell lymphoma (DLBCL) without central nervous system involvement. The cytokines of 13 healthy people and the lymphocyte subsets of 27 healthy people were measured as the control group. Results: We found a significant difference in the level of Th1/Th2/Th17 cytokines and lymphocyte subsets between PCNSL and healthy controls, especially IL-2, after treatment, which was significantly higher than before treatment (p<0.01). However, the level of CD19+ and CD4+/CD8+ decreased while CD8+ and CD3+ increased after treatment (regardless of whether the treatment was effective), and the difference was statistically significant. In addition, our analysis of different prognostic factors found that HD-MTX-based chemotherapy appears to have a longer progression-free survival and overall survival than osimertinib-based chemotherapy. Conclusion: There are significant differences in Th1/Th2/Th17 cytokines and lymphocyte subsets among PCNSL, DLBCL, and healthy controls, and their detection is helpful for the diagnosis, treatment, and prognosis of PCNSL. HD-MTX-based chemotherapy may still be the first choice for PCNSL.

Retracted: Targeting Transforming Growth Factor-Beta1 (TGF-ß1) Inhibits Tumorigenesis of Anaplastic Thyroid Carcinoma Cells Through ERK1/2-NFκB-PUMA Signaling.

The in vitro experiments of TGF-ß1 and the results of RT-PCR could not be repeated. In order not to affect others, the authors have asked for a retraction. Reference: Qiang Yin, Shan Liu, Anbing Dong, Xiufang Mi, Fengyun Hao, Kejun Zhang. Targeting Transforming Growth Factor-Beta1 (TGF-ß1) Inhibits Tumorigenesis of Anaplastic Thyroid Carcinoma Cells Through ERK1/2-NFkappakB-PUMA Signaling. Med Sci Monit 2016; 22: 2267-2277. DOI: 10.12659/MSM.898702.

Proteomic profiling identifies novel diagnostic biomarkers and molecular subtypes for mucinous tubular and spindle cell carcinoma of the kidney.

Mucinous tubular and spindle cell carcinoma (MTSCC) is a relatively rare renal epithelial neoplasm resembling type 1 papillary renal cell carcinoma (PRCC) morphologically and immunohistochemically. The accurate diagnosis of MTSCC remains a challenge. Here, by using proteomic profiling, we characterized MTSCC and PRCC to identify diagnostic biomarkers. We found that the MTSCC tumor proteome was significantly enriched in B-cell-mediated immunity when compared with the proteome of adjacent normal tissues of MTSCC or tumors of PRCC. Importantly, we identified MZB1, VCAN, and SOSTDC1 as diagnostic biomarkers to distinguish MTSCC from the solid variant of type 1 PRCC, with an AUC of 0.985 when combined. MZB1 was inversely correlated with tumor clinical stage and may play an anti-tumor role by activating the complement system. Finally, unsupervised clustering revealed two molecular subtypes of MTSCC, displaying different morphology, expression signatures of oxidative phosphorylation, and aggravation. In summary, our analyses identified a three-protein diagnostic panel and molecular subtypes for MTSCC. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Correlation of NUF2 Overexpression with Poorer Patient Survival in Multiple Cancers.

PURPOSE: NUF2 has been implicated in multiple cancers recently, suggesting NUF2 may play a role in the common tumorigenesis process. In this study, we aim to perform comprehensive meta-analysis of NUF2 expression in the cancer types included in the Cancer Genome Atlas (TCGA). MATERIALS AND METHODS: RNA-sequencing data in 31 cancer types in the TCGA data and 11 independent datasets were used to examine NUF2 expression. Silencing NUF2 using targeting shRNAs in hepatocellular carcinoma (HCC) cell lines was used to evaluate NUF2's role in HCC in vitro and in vivo. RESULTS: NUF2 up-regulation is significantly observed in 23 out of the 31 cancer types in the TCGA datasets and validated in 13 major cancer types using 11 independent datasets. NUF2 overexpression was clinically important as high NUF2 was significantly associated with tumor stages in eight different cancers. High NUF2 was also associated with significantly poorer patient overall survival and disease-free survival in eight and six cancers, respectively. We proceeded to validate NUF2 overexpression and its negative association with overall survival at the protein level in an independent cohort of 40 HCC patients. Compared to the non-targeting controls, NUF2 knockdown cells showed significantly reduced ability to grow, migrate into a scratch wound and invade the 8 µm porous membrane in vitro. Moreover, NUF2 knockdown cells also formed significantly smaller tumors than control cells in mouse xenograft assays in vivo. CONCLUSION: NUF2 up-regulation is a common feature of many cancers. The prognostic potential and functional impact of NUF2 up-regulation warrant further studies.

Immunoglobulin G4-related lymph node disease with an orbital mass mimicking Castleman disease: A case report.

BACKGROUND: Immunoglobulin (Ig) G4-associated diseases are a group of systemic diseases involving multiple organs and are also known as IgG4-associated sclerosing diseases. IgG4-associated lymphadenopathy occurring in the lymph nodes is characterized by a lack of specificity due to its clinicopathological characteristics and must be differentiated from a variety of lesions, such as Castleman disease, lymphatic follicular reactive hyperplasia, and lymphoma. CASE SUMMARY: A 65-year-old male patient, with Guillain-Barre syndrome for 5 years, presented to our hospital complaining of bilateral orbital mass for 2 years. After hospitalization, the results of the patient's laboratory tests showed that immunoglobulin subgroup IgG4 was 33.90 g/L and IgG was 30.30 g/L, but serum interleukin-6 was normal. The pathological morphology of orbital mass and cervical lymph node were consistent, which showed that a large number of plasma cells and eosinophils were observed in the lymphatic follicles, and the interstitial fibrous tissue was proliferative. Immunohistochemistry showed that CD20 (B cells) (+), CD3 (T cells) (+), CD38 (+), IgG (+), IgG4 positive cells > 100/high powered field, and IgG4/IgG > 40%. Combined with clinical and immunohistochemical results, lymphadenopathy was consistent with Castleman disease-like IgG4-associated sclerosing disease. Prednisone acetate treatment was given at 40 mg/d. After 2 wk, the superficial lymph nodes and orbital masses shrank, and the IgG4 level decreased. As prednisone acetate was regularly used at a reduced dosage, no recurrence of the disease has been observed. CONCLUSION: This case suggested that it is necessary to proceed cautiously in clinical practice with such patients, and immunoglobulin, complement, interleukin-6, C-reactive protein, and other examinations should be performed to confirm the diagnosis.

Sebaceous lymphadenocarcinoma of the parotid gland: A case report.

BACKGROUND: Sebaceous lymphadenoma is a benign tumor that occurs rarely in the salivary glands, most commonly in the parotid glands or periparotid lymph nodes, and even more rarely undergoes malignant transformation into a sebaceous lymphadenocarcinoma. CASE SUMMARY: We report an 82-year-old woman who presented with a painless mass in the right parotid region. We performed extended surgical resection of the parotid gland mass. Intraoperative pathology revealed a sebaceous lymphadenocarcinoma with metastasis into the periparotid cervical lymph nodes, so we also performed neck dissection and lymph node resection. Postoperative pathology confirmed the diagnosis. The literature review revealed that this was the seventh reported case of sebaceous lymphadenocarcinoma and the second reported case of cervical lymph node metastasis and infiltration of the skin of the parotid gland. CONCLUSION: Treatment of sebaceous lymphadenocarcinoma depends on the typing and clinical staging of the cancer. Extensive resection is the first choice, and adjuvant radiotherapy should be given to patients with high-grade tumors or those at an advanced clinical stage.

Clinical significance of eukaryotic translation initiation factor 5A2 in papillary thyroid cancer.

Eukaryotic translation initiation factor 5A2 (eIF5A2) plays an important role in tumor progression and prognosis evaluation. However, the potential role of eIF5A2 in human papillary thyroid cancer (PTC) is unknown. In this study, we aim to investigate the association between eIF5A2 expression and PTC clinical outcomes and underlying its Biological function in PTC cells in vitro and in vivo. The expression of eIF5A2 was examined by immunohistochemistry in PTC tissues and its adjacent tissue (n = 39) from 207 PTC patients. Functional analysis of eIF5A2 was performed in PTC cell lines in vitro and in vivo. The results showed that eIF5A2 was overexpressed in PTC tissues compared with the adjacent tissues. Enhanced eIF5A2 expression was significantly correlated with extrathyroidal extension (p = 0.012), lymph node metastasis (p = 0.002), TNM stage (p = 0.006), T classification (p = 0.047) and BRAF V600E mutation (p = 0.036). EIF5A2 inhibition prevented PTC cell growth, invasiveness and migration and induced cell apoptosis in vitro. Furthermore, eIF5A2 depletion inhibited tumor growth and metastasis in vivo. The data indicated that eIF5A2 could be employed as a novel prognostic marker and effective therapeutic target for PTC.

Regulation of GSK3ß/Nrf2 signaling pathway modulated erastin-induced ferroptosis in breast cancer.

Ferroptosis is a newly discovered form of regulated cell death and characterized by an iron-dependent accumulation of lethal lipid reactive oxygen species (ROS), ferroptosis may exhibit a novel spectrum of clinical activity for cancer therapy. However, the significance of ferroptosis in the context of carcinoma biology is still emerging. Glycogen synthase kinase-3ß (GSK-3ß) has been found to be a fundamental element in weaking antioxidant cell defense by adjusting the nuclear factor erythroid 2-related factor 2 (Nrf2). In our study, decreased expression of GSK-3ß was observed in the cancer tissues of breast cancer patients, results of immunohistochemistry indicated that Nrf2 was highly expressed in low-GSK-3ß-expressed breast cancer tissues. The contributions of aberrant expression of GSK-3ß and Nrf2 to the erastin-induced ferroptosis in breast cancer were further assessed, silence of GSK-3ß blocked erastin-induced ferroptosis with less production of ROS and malondialdehyde (MDA) via upregulation of GPX4 and downregulation of arachidonate 15-lipoxygenase (Alox15), overexpression of GSK-3ß enhanced erastin-triggered ferroptosis with elevated ROS and MDA. Enhanced erastin-induced ferroptosis by overexpression of GSK-3ß was blocked by activating Nrf2. We further confirmed that overexpression of GSK-3ß strengthened erastin-induced tumor growth inhibition in breast cancer xenograft models in vivo. In summary, our findings conclude that modulation the balance between GSK-3ß/Nrf2 is a promising therapeutic approach and probably will be important targets to enhance the effect of erastin-induced ferroptosis in breast cancer.

miR-199a-5p suppresses epithelial- mesenchymal-transition in anaplastic thyroid carcinoma cells via targeting Snail signals.

MicroRNAs (miRNAs) have been validated to play prominent roles in the occurrence and development of anaplastic thyroid carcinoma (ATC). miR-199a-5p was previously reported to act as a tumor suppressor or oncomiRNA in various types of cancer. However, its accurate expression, function, and mechanism in ATC remain unclear. Here, we find that miR-199a-5p is significantly downregulated in ATC tissues compared with adjacent non-cancerous tissues. Overexpression of miR-199a-5p significantly inhibits migration and invasion of ATC cells in vitro, and lung metastasis in vivo. Importantly, miR-199a-5p suppresses epithelial-mesenchymal transition (EMT) both in vitro and in vivo by targeting Snail. Taken together, this study reveals that miR-199a-5p is critical to the EMT progression in ATC cells. Targeting the pathway described here may be a novel approach for inhibiting metastasis of ATC.

miR-211 alleviates ischaemia/reperfusion-induced kidney injury by targeting TGFßR2/TGF-ß/SMAD3 pathway.

MicroRNA-211 (miR-211) is closely related to apoptosis and plays an important role in ischemia/reperfusion (I/R) injury. Whether miR-211 is involved in the protective effects in renal I/R injury is unknown. In this study, we evaluated the role of miR-211 in human tubular epithelial cells in response to hypoxia-reoxygenation (H/R) stimulation and I/R injury in vitro and in vivo. The results revealed that miR-211 was down-regulated and TGFßR2 was up-regulated in human kidney (HK-2) cells subjected to H/R. Luciferase reporter assay showed that TGFßR2 was a direct target of miR-211. Enforced miR-211 expression decreased H/R-induced HK-2 cell apoptosis and increased cell viability, and targeting miR-211 further increased H/R-induced HK-2 cell apoptosis and decreased cell viability. However, the effect of miR-211 was reversed by targeting TGFßR2 or enforced TGFßR2 expression in miR-211 overexpressing cells or miR-211 downexpressing cells. Moreover, we confirmed that miR-211 interacted with TGFßR2, and regulating TGF-ß/SMAD3 signal. In vivo in mice, miR-211 overexpression ameliorates biochemical and histological kidney injury, reduces apoptosis in mice following I/R. On the contrary, miR-211 downexpressing promoted histological kidney injury and increased apoptosis in mice following I/R. Inhibition of miR-211 or miR-211 overexpression inhibited TGF-ß/SMAD3 pathways or activated TGF-ß/SMAD3 signal pathways in vitro and in vivo, which are critical for cell survival. Our findings suggested that miR-211 suppress apoptosis and relieve kidney injury following H/R or I/R via targeting TGFßR2/TGF-ß/SMAD3 signals. Therefore, miR-211 may be as therapeutic potential for I/R- induced kidney injury.

EIF5A2 Is Highly Expressed in Anaplastic Thyroid Carcinoma and Is Associated With Tumor Growth by Modulating TGF- Signals.

Anaplastic thyroid carcinoma (ATC) is resistant to standard therapies and has no effective treatment. Eukaryotic translation initiation factor 5A2 (EIF5A2) has shown to be upregulated in many malignant tumors and proposed to be a critical gene involved in tumor metastasis. In this study, we aimed to investigate the expression status of EIF5A2 in human ATC tissues and to study the role and mechanisms of EIF5A2 in ATC tumorigenesis in vitro and in vivo. Expression of EIF5A2 protein was analyzed in paraffin-embedded human ATC tissues and adjacent nontumorous tissues (ANCT) (n=24) by immunochemistry. Expressions of EIF5A2 mRNA and protein were analyzed in fresh-matched ATC and ANCT (n=23) and ATC cell lines by real-time polymerase chain reaction (PCR) and Western blotting. The effect of targeting EIF5A2 with short hairpin RNA (shRNA) or EIF5A2 overexpression on the ATC tumorigenesis and TGF-/Smad2/3 signals in vitro and in vivo was investigated. Expression of EIF5A2 was significantly upregulated in ATC tissues and cell lines compared with ANCT and normal follicular epithelial cell line. Functional studies found that targeting EIF5A2 induced SW1736 cell death in vitro and in vivo, followed by significantly downregulated phosphorylation of Smad2/3 (p-Smad2/3) in SW1736 cells at the protein level. Ectopic expression of EIF5A2 could promote 8505C cell growth in vitro and in vivo, followed by significantly upregulated p-Smad3 at the protein level. Recombinant human TGF-1 (hTGF-1) treatment decreased the antiproliferative activity of the EIF5A2 downexpressing 8505C cells through reversing pSmad2/3. Using the specific inhibitor SB431542 to block TGF- pathway or Smad3 siRNA to knock down Smad3 increased the antiproliferative activity of the EIF5A2-overexpressing 8505C cells through inhibiting pSmad2/3. Our findings indicated that EIF5A2 controled cell growth in ATC cells, and EIF5A/TGF-/Smad2/3 signal may be a potential therapeutic target for ATC treatment.

miR-25-3p inhibition impairs tumorigenesis and invasion in gastric cancer cells in vitro and in vivo.

Deregulated expression of microRNAs (miRNAs) plays a role in the pathogenesis and progression of gastric cancer (GC). Among upregulated miRNAs, miR-25-3p has oncogenic potential and therefore represents an attractive target for the treatment of GC. Here, we investigated the role of miR-25-3p on GC cells in vitro and in vivo. We found that miR-25-3p overexpression significantly promoted growth and invasion of gastric cancer cells in vitro. Conversely, targeting miR-25-3p triggered significant inhibition of growth, invasion and migration in GC cells in vitro. In vivo delivery of miR-25-3p inhibitors induced significant anti-tumor activity in SCID mice bearing human GC xenografts. Our findings showed the evidence that in vivo antagonism of miR-25-3p impaired tumorigenesis, providing the rationale for clinical development of miR-25-3p inhibitors in GC.

Expression of long non-coding RNA H19 predicts distant metastasis in minimally invasive follicular thyroid carcinoma.

Downregulation of lncRNA H19 (H19) expression is associated with an unfavorable prognosis in some cancers. However, little was known as to whether there was an association between H19 and minimally invasive follicular thyroid carcinoma (MI-FTC). In our study, we used quantitative real-time polymerase chain reaction (qRT-PCR) to determine H19 expression in 186 patients with MI-FTC who underwent initial surgery. Of the 186 patients with MI-FTC, 21 patients show distant metastasis （M+）at the initial operation established the diagnosis of MI-FTC. Of the 165 patients who did not show distant metastasis at diagnosis during the follow-up period (≥10 years), 28 patients undergone M+ and 137 patients has no distant metastasis（M-）after the initial operation. Low H19 expression was associated with large tumor size, vascular invasion, and distant metastasis. Univariate analysis showed that gender (male), age (45 years or older), primary tumor size (4 cm or more), vascular invasion and H19 level (<1.12) were significant prognostic factors related to postoperative distant metastases. Multivariate analysis showed that age, primary tumor size (4 cm or more) and vascular invasion was a significant prognostic factor for survival. Patients with low H19 expression showed a poorer outcome in MI-FTC patients. Receiver-operating characteristic (ROC) curve analysis demonstrated H19 could distinguish M+ from M- patientswith a value of area under the curve (AUC). Our findings suggest that H19 is a potential prognostic factor for evaluating prognosis and the metastatic potential of MI-FTC at an initial operation stage.

S100A4 expression is associated with poor prognosis in patients with resectable gastrointestinal stromal tumor.

S100A4 is particularly associated with the progression and metastasis of numerous human malignancies. This study was designed to examine the clinicopathologic significance of S100A4 in gastrointestinal stromal tumor (GISTs). The level of OPNS100A4 expression in a large cohort of resectable GISTs was evaluated with immunohistochemistry. Its correlation with the clinicopathologic parameters of patients with resectable GISTs was analyzed. A survival analysis was performed to evaluate the prognostic significance of S100A4 expression using Kaplan-Meier method. Results: In 108 patients with resectable GISTs, the most high-risk GISTs had a strong level of S100A4 expression. Strong S100A4 expression was significantly associated with tumor size, mitosis, and recurrence, but not gender and age. Patients with weak S100A4 expression had a relatively longer disease-free survival compared to patients with strong S100A4 expression.Therefore, S100A4 expression is a putative marker for tumor progression and an adverse prognosis in GISTs.

Epigallocatechin gallate (EGCG) suppresses epithelial-Mesenchymal transition (EMT) and invasion in anaplastic thyroid carcinoma cells through blocking of TGF-ß1/Smad signaling pathways.

Transforming growth factor (TGF)-ß1 plays a crucial role in the epithelial-to-mesenchymal transition (EMT) in many cancer types and in thyroid cancers. Epigallocatechin-3-gallate (EGCG), the most important ingredient in the green tea, has been reported to possess antioxidant and anticancer activities. However, the cellular and molecular mechanisms explaining its action have not been completely understood. In this study, we found that EGCG significantly suppresses EMT, invasion and migration in anaplastic thyroid carcinoma (ATC) 8505C cells in vitro by regulating the TGF-ß/Smad signaling pathways. EGCG significantly inhibited TGF-ß1-induced expression of EMT markers (E-cadherin reduction and vimentin induction) in 8505C cells in vitro. Treatment with EGCG completely blocked the phosphorylation of Smad2/3, translocation of Smad4. Taken together, these results suggest that EGCG suppresses EMT and invasion and migration by blocking TGFß/Smad signaling pathways.

S100A4 Knockout Sensitizes Anaplastic Thyroid Carcinoma Cells Harboring BRAFV600E/Mt to Vemurafenib.

BACKGROUND/AIMS: Anaplastic thyroid cancer (ATC), with 25% BRAFV600E mutation, is one of the most lethal human malignancies that currently has no effective therapy. Vemurafenib, a BRAFV600E inhibitor, has shown promise in clinical trials, including ATC patients, but is being hampered by the acquisition of drug resistance. Therefore, combination therapy that includes BRAFV600E inhibition and avoids resistance is a clinical need. METHODS: ATC cell lines 8505C (BRAFV600E/mt), SW1736 (BRAFV600E/mt), KAT18 (BRAFV600E/wt) and Cal-62(BRAFV600E/wt) cells were used in the study. The ability of S100A knockout or /and in combination with the BRAF inhibitor vemurafenib on growth, apoptosis, invasion and apoptosis in ATC cells in vitro was demonstrated by MTT and BrdUrd incorporation assay, Annexin-V-FITC staining analyzed by flow cytometry, Transwell migration and Matrigel invasion assay. S100A4,pERK1/2, pAKT and pROCK1/2 protein was detected by western blot assay; Small molecule inhibitors of Y27632, U0126, MK-2206 and constitutively active forms of pCDNA-Myc-pERK, pCMV6-HA-Akt, pCMV-RhoA were employed, and the mechanistic studies were performed. We assessed the efficiency of in vivo combination treatment with S100A4 knockout and Vemurafenib on tumors. RESULTS: S100A4 knockout induced apoptosis and reduced proliferation by inactivation of pAKT and pERK signals, and inhibited invasion and migration by inactivation of pAKT and RhoA/ROCK1/2 signals in 8505C or Cal-62 cells in vitro, and vice versa in SW1736 and KAT18 cells. Vemurafenib did not affect apoptosis of both 8505C and SW1736 cells, but reduced proliferation via arresting cell cycle, and promoted cell migration and invasion in vitro. Combination treatment with S100A4 knockdown and vemurafenib reduced cell proliferation, migration and invasion in vitro compared to the S100A4 knockdown or Vemurafenib alone. Vemurafenib treatment resulted in a transient inhibition of pERK expression and gradually activation of pAKT expression, but quickly recovery from ERK1/2 activation inhibition by vemurafenib treatment in 4 h for SW1736 and 8505C cells. Combined treatment completely inhibited ERK1/2 and AKT activation during 48 h. In an in vivo mouse model of SW1736 and 8505C, vemurafenib treatment alone did not significantly inhibit tumor growth in both of the tumors, but inhibited tumor growth in combined groups. CONCLUSION: Our results show S100A4 knockout alone inhibits ATC cells (rich endogenous S100A4) survival and invasion, regardless of the BRAFV600E status, and potentiates the effect of vemurafenib on tumor regression in vitro and in vivo. In addition, S100A4 knockout potently inhibits the recovery from ERK1/2 activation inhibition and the AKT activation following vemurafenib treatment and reversed the vemurafenib resistance. This therapeutic combination may be of benefit in patients with ATC.

Clusterin Reduces Cold Ischemia-Reperfusion Injury in Heart Transplantation Through Regulation of NF-kB Signaling and Bax/Bcl-xL Expression.

BACKGROUND/AIMS: Ischemia-reperfusion (I/R) injury is an unavoidable event occurring during heart transplantation and is a key factor in graft failure and the long-term survival rate of recipients. Therefore, there is an urgent need for the development of new therapies to prevent I/R injury. Clusterin is a hetero-dimeric glycoprotein with an antiapoptotic function. In this study, we investigated whether clusterin was cardioprotective in heart transplantation against I/R injury using an in vivo rat model and an in vitro cell culture system, and examined the underlying mechanisms of I/R injury. METHODS: Heart grafts from wild-type C57BL/6 mice were preserved in UW solution (control) or UW solution containing recombinant human apolipoprotein-J (hr clusterin) for 24 h. The preserved hearts were implanted into recipient mice of the same strain as the donors for 72 h, and the heart grafts were then taken for histopathological and gene expression analyses. An in vitro ischemia reperfusion model using H9C2 cells or H9C2/clusterin cDNA cells was constructed. The expression of clusterin, p65, Bax, Bcl-xL, IL-1ß, and TNF-α protein and mRNA in heart tissue and H9C2 cells was detected by western blot, reverse transcription-polymerase chain reaction (RT-PCR), and quantitative RT-PCR assays; IL-1ß and TNF-α protein was detected by enzyme-linked immunosorbent assays; NF-kB activity was detected by an electrophoretic mobility shift assay; cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and flow cytometric analyses. RESULTS: Cold I/R caused severe morphologic myocardial injury to heart grafts from wild-type C57BL/6 mice, whereas grafts from hr clusterin preservation showed less damage, as demonstrated by decreased cell apoptosis/death, decreased neutrophil infiltration, and the preservation of the normal structure of the heart. Clusterin reduced the expression of p65, pre-inflammatory IL-1ß, and TNF-α, and the pro-apoptotic gene Bax, while it enhanced the expression of the anti-apoptotic gene Bcl-xL in vitro and in vivo. Clusterin inhibited cell apoptosis/death and reduced pre-inflammatory. CONCLUSION: Clusterin is a promising target for preventing cold I/R injury in heart transplantation. This study also shows that the resultant protective effects of clusterin are mediated by NF-κB signaling and Bax/Bcl-xL expression.