Construction and validation of risk model of EMT-related prognostic genes for kidney renal clear cell carcinoma.

BACKGROUND: Kidney renal clear cell carcinoma (KIRC) is a prevalent type of urological malignancy. The present study aimed to predict biomarkers for KIRC. METHODS: We collected transcriptomic and clinical information for KIRC from The Cancer Genome Atlas and GSE22541 cohorts. RESULTS: Unsupervised clustering of 35 epithelial-mesenchymal transformation (EMT)-related differentially expressed gene profiles divided samples into two clusters with distinct immune characteristics. Six genes (IL20RB, DDC, ANKRD36BP2, F2RL1, TEK, and AMN) were found to construct a prognostic risk model using multivariate Cox regression analysis. Kaplan-Meier analysis suggested the better prognosis of the KIRC patients in the low-risk group than that in the high-risk group. Immune infiltration analyses was conducted using xCell and single-sample gene set enrichment analysis, indicating that the risk score was associated with the immune microenvironment of the KIRC. Prognostic marker gene-targeted medications with high drug sensitivity were predicted in KIRC patients. CONCLUSIONS: In summary, the present study identified IL20RB, DDC, ANKRD36BP2, F2RL1, TEK, and AMN as prognostic biomarkers, providing insight into immunotherapy and gene-targeted drugs of KIRC.

Clearance of serum solutes by hemofiltration in dogs with severe heat stroke.

BACKGROUND: We have previously reported that hemofiltration (HF) may be an effective additional means of treating heat stroke when rapid cooling is not effective. METHODS: Dogs were assigned to a heat stroke (control) or heat stroke + hemofiltration (HF) group (n = 8 each group). After heat stroke induction, dogs in the HF group received HF for 3 h. Serum concentrations of interleukin (IL)-10, tumor necrosis factor (TNF)-α, IL-6, blood urea nitrogen (BUN) and creatinine were measured at baseline and 1, 2, and 3 h after heat stroke. Clearance rates of solutes were determined 1, 2, and 3 h after the start of HF. RESULTS: Serum concentrations of all solutes tended to increase with time after heat stroke in the control group, but decreased (BUN, creatinine) or remained relatively unchanged (TNF-α, IL-6, IL-10) with time in the HF group. Concentrations of all solutes were significantly lower in the HF group compared with the control group at 2 and 3 h (P < 0.05). Clearance rates for small molecular weight solutes were high, while those for larger molecular weight solutes were low. CONCLUSION: HF prevents heat stroke-induced increases in serum cytokine concentrations and is effective for clearing small molecular weight solutes from serum, but less effective for clearing larger molecular weight solutes, including TNF-α, IL-6, and IL-10.

The effects of genistein on transforming growth factor-ß1-induced invasion and metastasis in human pancreatic cancer cell line Panc-1 in vitro.

BACKGROUND: Pancreatic cancer is a devastating disease with the worst mortality rate. Therefore, a rational strategy for future drug development is critical. Genistein is a small, biologically active flavonoid that is found in high amounts in soy. This important compound supports a wide variety of biological activities, but is best known for its ability to inhibit cancer progression. METHODS: Transwell chamber assay was performed to determine the effect of genistein on the invasion of the human pancreatic cancer cell line Panc-1 induced by transforming growth factor-ß1 (TGF-b1) in the different condition (5 ng/ml 24 hours and 10 ng/ml 48 hours); Reverse transcription-polymerase chain reaction (RT-PCR) was used to estimate the mRNA levels of urinary plasminogen activator (uPA), matrix metallopeptidase 2/9 (MMP-2/9), Smad4, E-Cadherin and Vimentin; Western blotting was used to detect the protein levels of uPA, E-Cadherin, ERK1/2, P38 and P-P38, and the activity of MMP-2/9 protein were detected by gelatin zymography assay method. Cells structure was observed and analyzed by microscopy. RESULTS: Genistein can inhibit effectively TGF-b1-induced invasion and metastasis in Panc-1 by Transwell assay, which is through regulating the mRNA and protein expression of uPA and MMP2, but not MMP9 by RT-PCR/Western blotting, and is positively correlated with the concentration of genistein. At the same time, genistein also could improve the progress of epithelial-mesenchymal transition (EMT) via morphology observation using light microscopy/transmission electron microscopy (TEM), which is mediated by the down-regulation of E-cadherin and the up-regulation of vimentin. CONCLUSIONS: TGF-b1 mediates EMT process via numerous intracellular signal transduction pathways. The potential molecular mechanisms are all or partly through Smad4-dependent and -independent pathways (p38 MAPK) to regulate the antitumor effect of genistein.

Preliminary study on the freeze-drying of human bone marrow-derived mesenchymal stem cells.

Long-term preservation and easy transportation of human bone marrow-derived mesenchymal stem cells (hBM-MSCs) will facilitate their application in medical treatment and bioengineering. A pilot study on the freeze-drying of hBM-MSCs was carried out. hBM-MSCs were loaded with trehalose. The glass transition temperature of the freeze-drying suspension was measured to provide information for the cooling and primary drying experiment. After freeze-drying, various rehydration processes were tested. The highest recovery rate of hBM-MSCs was (69.33±13.08)%. Possible methods to improve freeze-drying outcomes are discussed. In conclusion, the present study has laid a foundation for the freeze-drying hBM-MSCs.

[Preparation of a novel telomerase inhibitory protein LPTS-L].

The gene for LPTS is originally cloned as a human liver-related putative tumor suppressor (LPTS) gene that encodes a full length protein of 328 amino acids (LPTS-L). LPTS-L is also identified as a telomerase inhibitor to regulate telomere length in the cells. To facilitate the functional and structural studies of LPTS-L protein, the cDNA for LPTS-L was cloned into the expression vector pET-24 in frame to generate a recombinant plasmid pET-24-LPTS. The LPTS-L protein was expressed in E. coli BL21 solublely, and purified by Ni Sepharose affinity chromatography which, however, is not fit for large scale protein purification. The gene of LITS-L was then PCR amplified to remove the 6 x His tag, and cloned into pET-24a. The non-fusion protein of LPTS-L was expressed in E. coli B21, and purified by phosphocellulose P11 chromatography. The purity of LPTS-L protein was about 55% after that procedure,and arrived at 80% after second purification by Sephadex G-100 chromatography. Western Blotting analysis showed that the band reflects the specific binding of anti-LPTS antiserum against the purified LPTS-L protein. The TRAP assay was performed to detect the telomerase inhibitory activity of LPTS-L protein in vitro. It was observed that the purified LPTS-L inhibited the activity of telomerase greatly, similarly with that of LPTS-L protein purified by Ni Sepharose 4B. Our results suggest that phosphocellulose P11 plus Sephadex G-100 chromatography could substitute for Ni Sepharose 4B affinity chromatography for preparation of purified LPTS-L protein. Through this study, a technique for preparation of LPTS-L protein in a large scale is established.

[Treatment of hemolytic uremic syndrome after acute stage].

OBJECTIVE: Hemolytic uremic syndrome (HUS) is a common primary disease that can cause acute renal failure in childhood. Renal disease is the most important long-term complication in patients who survived the acute stage of HUS. Use of angiotensin-converting enzyme inhibitors (ACEI) and a restricted protein intake may be beneficial to the patients. However, it is not established whether such patients should be treated with steroids and immunosuppressors. The present study aimed to probe into the benefit of using steroid and immunosuppressor in patients after acute stage of HUS. METHODS: The subjects included 17 patients (aged 9 months to 15 years, 12 males, 5 females) with HUS. Thirteen patients recovered from the acute stage of HUS, and underwent continuative treatment and follow-up. All the patients were treated with ACEI and early restriction of protein intake. Additionally, 2 children manifested as glomerulonephritis, one was treated with triperygium glycosides. Other 11 children who manifested as nephrotic syndrome were treated with prednisone, among them 5 children had no response or had incomplete response to prednisone, for these children short-term high dose cyclophosphamide or methylprednisolone pulse treatment were added; in 3 of the children short-term high dose methylprednisolone treatment was applied additionally for membranoproliferative glomerulonephritis and/or focal segmental glomerulosclerosis and crescentic glomerulonephritis. RESULTS: After follow-up for 2 months to 8 years, 4 patients with milder disease recovered, their blood pressure, renal function and urinalysis became normal, but 1 patient had recurrence. Among 9 patients with severe disease, 6 maintained normal blood pressure, recovered renal function and urinalysis, the other 3 patients failed to comply with treatment protocol and died during the 3rd, 9th and 13th month. The remainder (4 cases) gave up therapy and died on the 27th to 48th days of the course. CONCLUSION: The treatment applied in this study could improve the prognosis of patients after acute phase of HUS evidently by using the steroid and immuno suppressor according to clinical classification and pathological findings. It is recommended that triperygium glycosides is beneficial to children with glomerulonephritis, proteinuria and hematuria after acute stage of HUS. Adjustment of therapeutic schedule based on pathological findings after renal biopsy is helpful. To the patients with progressive renal failure who have no response to the steroid and immunosuppressors, steroid and immunosuppressor should be discontinued and dialysis treatment should be applied. Protocol compliance is also an important factor.