Tamoxifen ameliorates obstructive nephropathy through Src and the PI3K/Akt/mTOR pathway.

BACKGROUND INFORMATION: Tubulointerstitial fibrosis is the end-point of chronic kidney diseases. Tamoxifen, a selective oestrogen receptor (ER) modulator, attenuates renal fibrosis, by regulating the transforming growth factor (TGF)-ß/Smad signalling. Src and phosphoinositide 3-kinase (PI3K)/Akt pathways play critical roles in the pathogenesis of renal fibrosis. However, the activation of the non-canonical TGF-ß signalling in renal fibrosis after treatment with tamoxifen remains unclear. Renal fibrosis was induced by unilateral ureteral obstruction (UUO) in rats. Tamoxifen was orally administered after UUO. Additionally, HK-2 cells were treated with tamoxifen in the presence or absence of TGF-ß1. The selective ER down-regulator ICI and ER-α silencing were used to confirm the involvement of ER-α on the effect of tamoxifen on TGF-ß1-stimulated fibrosis in HK-2 cells. RESULTS: Tamoxifen treatment ameliorated UUO-induced renal fibrosis as shown by decreased expression of α-smooth muscle actin (SMA), fibronectin and connective tissue growth factor (CTGF). The phosphorylation of Src, PI3K, Akt, mammalian target of rapamycin (mTOR) and p70S6K significantly decreased in UUO kidneys from tamoxifen-treated animals. Tamoxifen dose-dependently suppressed the TGF-ß1-induced expression of α-SMA and CTGF, and phosphorylation of Src, PI3K, Akt, mTOR and p70S6K in HK-2 cells. These anti-fibrotic effects were reversed by treatment with ICI and silencing of ER-α. Moreover, inhibition of the PI3K/Akt and mTOR/p70S6K pathways was observed in HK-2 cells co-treated with PP1 (a Src kinase inhibitor) and tamoxifen. CONCLUSIONS: The anti-fibrotic effects of tamoxifen are associated with the suppression of Src kinase function via ER-α, followed by inhibition of the PI3K/Akt and mTOR/p70S6K signalling pathways. SIGNIFICANCE: Our findings suggest that tamoxifen is a novel therapeutic option for the prevention and treatment of renal fibrosis.

Janus kinase 2 inhibition by Licochalcone B suppresses esophageal squamous cell carcinoma growth.

Esophageal cancer (EC) is one of the leading causes to cancer death in the worldwide and major population of EC is esophageal squamous cell carcinoma (ESCC). Still, ESCC-targeted therapy has not been covered yet. In the present study we have identified that Licochalcone B (Lico B) inhibited the ESCC growth by directly blocking the Janus kinase (JAK) 2 activity and its downstream signaling pathway. Lico B suppressed KYSE450 and KYSE510 ESCC cell growth, arrested cell cycle at G2/M phase and induced apoptosis. Direct target of Lico B was identified by kinase assay and verified with in vitro and ex vivo binding. Computational docking model predicted for Lico B interaction to ATP-binding pocket of JAK2. Furthermore, treatment of JAK2 clinical medicine AZD1480 to ESCC cells showed similar tendency with Lico B. Thus, JAK2 downstream signaling proteins phosphorylation of STAT3 at Y705 and S727 as well as STAT3 target protein Mcl-1 expression was decreased with treatment of Lico B. Our results suggest that Lico B inhibits ESCC cell growth, arrests cell cycle and induces apoptosis, revealing the underlying mechanism involved in JAK2/STAT3 signaling pathways after Lico B treatment. It might provide potential role of Lico B in the treatment of ESCC.

Dual inhibition of EGFR and MET by Echinatin retards cell growth and induces apoptosis of lung cancer cells sensitive or resistant to gefitinib.

Patients with non-small-cell lung cancer (NSCLC) containing epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to tyrosine kinase inhibitor gefitinib; however, the treatment is less effective over time. Gefitinib resistance mechanisms include MET gene amplification. A therapeutic strategy targeting MET as well as EGFR can overcome resistance to gefitinib. In the present study we identified Echinatin (Ecn), a characteristic chalcone in licorice, which inhibited both EGFR and MET and strongly altered NSCLC cell growth. The antitumor efficacy of Ecn against gefitinib-sensitive or -resistant NSCLC cells with EGFR mutations and MET amplification was confirmed by suppressing cell proliferation and anchorage-independent colony growth. During the targeting of EGFR and MET, Ecn significantly blocked the kinase activity, which was validated with competitive ATP binding. Inhibition of EGFR and MET by Ecn decreases the phosphorylation of downstream target proteins ERBB3, AKT and ERK compared with total protein expression or control. Ecn induced the G2/M cell cycle arrest, and apoptosis via the intrinsic pathway of caspase-dependent activation. Ecn induced ROS production and GRP78, CHOP, DR5 and DR4 expression as well as depolarized the mitochondria membrane potential. Therefore, our results suggest that Ecn is a promising therapeutic agent in NSCLC therapy.

Deoxypodophyllotoxin, a Lignan from Anthriscus sylvestris, Induces Apoptosis and Cell Cycle Arrest by Inhibiting the EGFR Signaling Pathways in Esophageal Squamous Cell Carcinoma Cells.

Deoxypodophyllotoxin (DPT) derived from Anthriscus sylvestris (L.) Hoffm has attracted considerable interest in recent years because of its anti-inflammatory, antitumor, and antiviral activity. However, the mechanisms underlying DPT mediated antitumor activity have yet to be fully elucidated in esophageal squamous cell carcinoma (ESCC). We show here that DPT inhibited the kinase activity of epidermal growth factor receptor (EGFR) directly, as well as phosphorylation of its downstream signaling kinases, AKT, GSK-3ß, and ERK. We confirmed a direct interaction between DPT and EGFR by pull-down assay using DPT-beads. DPT treatment suppressed ESCC cell viability and colony formation in a time- and dose-dependent manner, as shown by MTT analysis and soft agar assay. DPT also down-regulated cyclin B1 and cdc2 expression to induce G2/M phase arrest of the cell cycle and upregulated p21 and p27 expression. DPT treatment of ESCC cells triggered the release of cytochrome c via loss of mitochondrial membrane potential, thereby inducing apoptosis by upregulation of related proteins. In addition, treatment of KYSE 30 and KYSE 450 cells with DPT increased endoplasmic reticulum stress, reactive oxygen species generation, and multi-caspase activation. Consequently, our results suggest that DPT has the potential to become a new anticancer therapeutic by inhibiting EGFR mediated AKT/ERK signaling pathway in ESCC.

Isolinderalactone Induces Cell Death via Mitochondrial Superoxide- and STAT3-Mediated Pathways in Human Ovarian Cancer Cells.

The mortality rate of ovarian cancer (OC) worldwide increases with age. OC is an often fatal cancer with a curative rate of only 20-30%, as symptoms often appear after disease progression. Studies have reported that isolinderalactone (ILL), a furanosesquiterpene derivative extracted from the dried root of Lindera aggregata, can inhibit several cancer cell lines' growth. However, the molecular mechanisms underlying ILL activities in human OC cells remain unexplored. This study investigated the antitumor activities of ILL in human OC cells by inducing mitochondrial superoxide (mtSO) and JAK-signal transducer and activator of transcription 3 (STAT3)-dependent cell death. ILL caused cell death in SKOV-3 and OVCAR-3 cells and increased the cell proportion in the subG1 phase. Additionally, ILL significantly induced mtSO production and reduced ROS production. Moreover, ILL downregulated mitochondrial membrane potential and the expression levels of anti-apoptotic Bcl-2 family proteins and superoxide dismutase (SOD)2. Results showed that ILL decreased phosphorylation of serine 727 and tyrosine 705 of STAT3 and expression of survivin, a STAT3-regulated gene. Furthermore, ILL-induced cell death was reversed by pretreatment of Mito-TEMPO, a mitochondria-specific antioxidant. These results suggest that ILL induces cell death by upregulation of mtSO, downregulation of mitochondrial SOD2, and inactivation of the STAT3-mediated pathway.

MED28 Over-Expression Shortens the Cell Cycle and Induces Genomic Instability.

The mammalian mediator complex subunit 28 (MED28) is overexpressed in a variety of cancers and it regulates cell migration/invasion and epithelial-mesenchymal transition. However, transcription factors that increase MED28 expression have not yet been identified. In this study, we performed a luciferase reporter assay to identify and characterize the prospective transcription factors, namely E2F transcription factor 1, nuclear respiratory factor 1, E-26 transforming sequence 1, and CCAAT/enhancer-binding protein ß, which increased MED28 expression. In addition, the release from the arrest at the G1-S or G2-M phase transition after cell cycle synchronization using thymidine or nocodazole, respectively, showed enhanced MED28 expression at the G1-S transition and mitosis. Furthermore, the overexpression of MED28 significantly decreased the duration of interphase and mitosis. Conversely, a knockdown of MED28 using si-RNA increased the duration of interphase and mitosis. Of note, the overexpression of MED28 significantly increased micronucleus and nuclear budding in HeLa cells. In addition, flow cytometry and fluorescence microscopy analyses showed that the overexpression of MED28 significantly increased aneuploid cells. Taken together, these results suggest that MED28 expression is increased by oncogenic transcription factors and its overexpression disturbs the cell cycle, which results in genomic instability and aneuploidy.

Retrochalcone Echinatin Triggers Apoptosis of Esophageal Squamous Cell Carcinoma via ROS- and ER Stress-Mediated Signaling Pathways.

Esophageal squamous cell carcinoma (ESCC) is a poor prognostic cancer with a low five-year survival rate. Echinatin (Ech) is a retrochalone from licorice. It has been used as a herbal medicine due to its anti-inflammatory and anti-oxidative effects. However, its anticancer activity or underlying mechanism has not been elucidated yet. Thus, the objective of this study was to investigate the anti-tumor activity of Ech on ESCC by inducing ROS and ER stress dependent apoptosis. Ech inhibited ESCC cell growth in anchorage-dependent and independent analysis. Treatment with Ech induced G2/M phase of cell cycle and apoptosis of ESCC cells. It also regulated their related protein markers including p21, p27, cyclin B1, and cdc2. Ech also led to phosphorylation of JNK and p38. Regarding ROS and ER stress formation associated with apoptosis, we found that Ech increased ROS production, whereas its increase was diminished by NAC treatment. In addition, ER stress proteins were induced by treatment with Ech. Moreover, Ech enhanced MMP dysfunction and caspases activity. Furthermore, it regulated related biomarkers. Taken together, our results suggest that Ech can induce apoptosis in human ESCC cells via ROS/ER stress generation and p38 MAPK/JNK activation.

MMP11 and CD2 as novel prognostic factors in hormone receptor-negative, HER2-positive breast cancer.

PURPOSE: More accurate prediction of patient outcome based on molecular subtype is required to identify patients who will benefit from specific treatments. METHODS: We selected novel 16 candidate prognostic genes, including 10 proliferation-related genes (p-genes) and 6 immune response-related genes (i-genes), from the gene list identified in our previous study. We then analyzed the association between their expression, measured by quantitative real-time reverse transcription-PCR in formalin-fixed, paraffin-embedded tissues, and clinical outcome in 819 breast cancer patients according to molecular subtype. RESULTS: The prognostic significance of clinical and gene variables varied according to the molecular subtype. Univariate analysis showed that positive lymph node status was significantly correlated with the increased risk of distant metastasis in all subtypes except the hormone receptor-negative, HER2-positive (HR-/HER2+) subtype. Most p-genes were significantly associated with poor prognosis in patients with the HR+/HER2- subtype, whereas i-genes correlated with a favorable outcome in patients with HR-/HER2+ breast cancer. In HR-/HER2+ breast cancer, four genes (three i-genes BTN3A2, CD2, and TRBC1 and the p-gene MMP11) were significantly associated with distant metastasis-free survival (DMFS). A new prognostic model for HR-/HER2+ breast cancer based on the expression of MMP11 and CD2 was developed and the DMFS for patients in the high-risk group according to our model was significantly lower than that for those in the low-risk group. Multivariate analyses revealed that our risk score is an independent prognostic factor for DMFS. Moreover, C-index showed that our risk score has a superior prognostic performance to traditional clinicopathological factors. CONCLUSIONS: Our new prognostic model for HR-/HER2+ breast cancer provides more accurate information on the risk of distant metastasis than traditional clinical prognostic factors and may be used to identify patients with a good prognosis in this aggressive subtype of breast cancer.

Identification of cromolyn sodium as an anti-fibrotic agent targeting both hepatocytes and hepatic stellate cells.

Liver fibrosis and cirrhosis, the late stage of fibrosis, are threatening diseases that lead to liver failure and patient death. Although aberrantly activated hepatic stellate cells (HSCs) are the main cause of disease initiation, the symptoms are primarily related to damaged hepatocytes. Thus, damaged hepatocytes, as well as HSCs, need to be simultaneously considered as therapeutic targets to develop more efficient treatments. Here, we suggest cromolyn sodium as an anti-fibrotic agent to commonly modulate hepatocytes and hepatic stellate cells. The differentially expressed genes from 6 normal and 40 cirrhotic liver tissues which were collected from GEO data were assessed by pharmacokinetic analysis using a connectivity map to identify agents that commonly revert abnormal hepatocytes and HSCs to normal conditions. Based on a series of analyses, a few candidates were selected. Candidates were tested in vitro to determine their anti-fibrotic efficacy on HSCs and hepatocytes. Cromolyn, which was originally developed as a mast cell stabilizer, showed the potential to ameliorate activated HSCs in vitro. The activation and collagen accumulation for HSC cell lines LX2 and HSC-T6 were reduced by 50% after cromolyn treatment at a low concentration without apoptosis. Furthermore, cromolyn treatment compromised the TGF-ß-induced epithelial mesenchyme transition and replicative senescence rate of hepatocytes, which are generally associated with fibrogenesis. Taken together, cromolyn may be the basis for an effective cure for fibrosis and cirrhosis because it targets both HSCs and hepatocytes.

Alpha-lipoic acid attenuates lipopolysaccharide-induced kidney injury.

BACKGROUND: Kidney is one of the major target organs in sepsis, while effective prevention of septic acute kidney injury has not yet been established. α-Lipoic acid (LA) has been known to exert beneficial effects against lipopolysaccharide (LPS)-induced damages in various organs such as heart, lung, and liver. We investigated the protective effect of LA on LPS-induced kidney injury. METHODS: Two groups of rats were treated with LPS (20 mg/kg, i.p.), one of which being co-treated with LA (50 mg/kg), while the control group was treated with vehicle alone. Human renal proximal tubular epithelial cells (HK-2 cells) were cultured with or without LPS (10 µg/ml) in the presence or absence of LA (100 µg/ml) for 3 h prior to LPS treatment. RESULTS: Serum creatinine level was increased in LPS-treated rats, which was attenuated by LA co-treatment. LPS treatment induced cleaved caspase-3 expression in the kidney, which was counteracted by LA. Terminal deoxynucleotidyl transferase dUTP nick-end labeling-positive cells increased in the kidneys of LPS-treated rats compared with controls, which was counteracted by LA treatment. Protein expression of inducible nitric oxide synthase and cyclooxygenase-2 detected by immunoblotting and/or immunohistochemical staining, along with mRNA levels of pro-inflammatory cytokines detected by real-time polymerase chain reaction, was increased in the kidney with LPS administration, which was ameliorated with LA treatment. LA also protected LPS-induced tubular dysfunction, preserving type 3 Na(+)/H(+) exchanger and aquaporin 2 expressions in the kidney. Suppression of LPS-induced expression of cleaved caspase-3 by LA was also observed in HK-2 cells. Increased protein expression of phospho-extracellular signal-regulated kinases 1/2 and c-Jun N-terminal kinases by LPS treatment was attenuated by LA pretreatment, while p38 was not affected by either LPS or LA treatment. MitoTracker Red demonstrated LA prevented LPS-induced increment of mitochondrial oxidative stress, where concurrent 4',6-diamidino-2-phenylindole staining also revealed marked fragmentation and condensation of nuclei in HK-2 cells treated with LPS, which was prevented by LA. CONCLUSION: LA treatment attenuates LPS-induced kidney injury, such as renal tubular dysfunction, by suppression of apoptosis, and inflammation.

Fluoxetine Decreases the Proliferation and Adipogenic Differentiation of Human Adipose-Derived Stem Cells.

Fluoxetine was originally developed as an antidepressant, but it has also been used to treat obesity. Although the anti-appetite effect of fluoxetine is well-documented, its potential effects on human adipose-derived stem cells (ASCs) or mature adipocytes have not been investigated. Therefore, we investigated the mechanisms underlying the inhibitory effects of fluoxetine on the proliferation of ASCs. We also investigated its inhibitory effect on adipogenic differentiation. Fluoxetine significantly decreased ASC proliferation, and signal transduction PCR array analysis showed that it increased expression of autophagy-related genes. In addition, fluoxetine up-regulated SQSTM1 and LC3B protein expression as detected by western blotting and immunofluorescence. The autophagy inhibitor, 3-methyladenine (3-MA), significantly attenuated fluoxetine-mediated effects on ASC proliferation and SQSTM1/LC3B expression. In addition, 3-MA decreased the mRNA expression of two autophagy-related genes, beclin-1 and Atg7, in ASCs. Fluoxetine also significantly inhibited lipid accumulation and down-regulated the levels of PPAR-γ and C/EBP-α in ASCs. Collectively, these results indicate that fluoxetine decreases ASC proliferation and adipogenic differentiation. This is the first in vitro evidence that fluoxetine can reduce fat accumulation by inhibiting ASC proliferation and differentiation.

The Photoprotective Effect of S-Methylmethionine Sulfonium in Skin.

S-Methylmethionine sulfonium (SMMS) was reported to have wound-healing effects; we therefore have investigated the photoprotective effect of SMMS in the present study. SMMS increased the viability of keratinocyte progenitor cells (KPCs) and human dermal fibroblasts (hDFs) following ultraviolet B (UVB) irradiation, and reduced the UVB-induced apoptosis in these cells. SMMS increased the phosphorylation of extracellular signal-regulated kinases (ERK), and the inhibitor of the mitogen-activated protein kinase pathway significantly decreased the SMMS-induced viability of KPCs and hDFs. In addition, SMMS attenuated the UVB-induced reactive oxygen species (ROS) generation in KPCs and hDFs. SMMS induced the collagen synthesis and reduced the matrix metalloproteinase-1 expression in UVB-irradiated hDFs. In animal studies, application of 5% and 10% SMMS before and after UVB-irradiation significantly decreased the UVB-induced erythema index and depletion of Langerhans cells. In summary, SMMS protects KPCs and hDFs from UVB irradiation, and reduces UVB-induced skin erythema and immune suppression. Therefore, SMMS can be used as a cosmetic raw material, and protect skin from UVB.

Domperidone, a Dopamine Receptor D2 Antagonist, Induces Apoptosis by Inhibiting the ERK/STAT3-Mediated Pathway in Human Colon Cancer HCT116 Cells.

Colorectal cancer (CRC) continues to demonstrate high incidence and mortality rates, emphasizing that implementing strategic measures for prevention and treatment is crucial. Recently, the dopamine receptor D2 (DRD2), a G protein-coupled receptor, has been reported to play multiple roles in growth of tumor cells. This study investigated the anticancer potential of domperidone, a dopamine receptor D2 antagonist, in HCT116 human CRC cells. Domperidone demonstrated concentration- and time-dependent reductions in cell viability, thereby inducing apoptosis. The molecular mechanism revealed that domperidone modulated the mitochondrial pathway, decreasing mitochondrial Bcl-2 levels, elevating cytosolic cytochrome C expression, and triggering caspase- 3, -7, and -9 cleavage. Domperidone decreased in formation of ß-arrestin2/MEK complex, which contributing to inhibition of ERK activation. Additionally, treatment with domperidone diminished JAK2 and STAT3 activation. Treatment of U0126, the MEK inhibitor, resulted in reduced phosphorylation of MEK, ERK, and STAT3 without alteration of JAK2 activation, indicating that domperidone targeted both MEK-ERK-STAT3 and JAK2-STAT3 signaling pathways, respectively. Immunoblot analysis revealed that domperidone also downregulated DRD2 expression. Domperidone-induced reactive oxygen species (ROS) generation and N-acetylcysteine treatment mitigated ROS levels and restored cell viability. An in vivo xenograft study verified the significant antitumor effects of domperidone. These results emphasize the multifaceted anticancer effects of domperidone, highlighting its potential as a promising therapeutic agent for human CRC.

EP4 receptor agonist CAY10598 upregulates ROS-dependent Hsp90 cleavage in colorectal cancer cells.

Prostaglandin E2 (PGE2) interacts with four specific G protein-coupled receptors, namely EP1, EP2, EP3, and EP4, playing a pivotal role in determining the fate of cells. Our previous findings highlighted that stimulating the EP4 receptor with its agonist, CAY10598, triggers apoptosis in colon cancer HCT116 cells via the production of reactive oxygen species (ROS). This process also reduces the phosphorylation of the oncogenic protein JAK2 and leads to its degradation in these cells. In this study, our goal was to explore the pathways through which CAY10598 leads to JAK2 degradation. We focused on Hsp90, a heat shock protein family member known for its role as a molecular chaperone maintaining the stability of several key proteins including EGFR, MET, Akt, and JAK2. Our results show that CAY10598 decreases the levels of client proteins of Hsp90 in HCT116 cells, an effect reversible by pretreatment with the ROS scavenger N-acetyl cysteine (NAC) or the proteasome inhibitor MG132, indicating that the degradation is likely driven by ROS. Furthermore, we observed that CAY10598 cleaves both α and ß isoforms of Hsp90, the process inhibited by NAC. Inhibition of EP4 with the antagonist GW627368x not only prevented the degradation of Hsp90 client proteins but also the cleavage of Hsp90 itself in CAY10598-treated HCT116 cells. Additionally, CAY10598 suppressed the growth of HCT116 cells implanted in mice. Our findings reveal that CAY10598 induces apoptosis in cancer cells by a novel mechanism involving the ROS-dependent cleavage of Hsp90, thereby inhibiting the function of crucial Hsp90 client proteins.

Translation Initiation Factor-2S2 (eIF2S2) Contributes to Cervical Carcinogenesis by Inhibiting the TGF-ß/SMAD4 Signaling Pathway.

The deregulation of protein translational machinery and the oncogenic role of several translation initiation factors have been extensively investigated. This study aimed to investigate the role of eukaryotic translation initiation factor 2S2 (eIF2S2, also known as eIF2ß) in cervical carcinogenesis. Immunohistochemical analysis of human cervical carcinoma tissues revealed a stage-specific increase in eIF2S2 expression. The knockdown of eIF2S2 in human cervical cancer (SiHa) cells significantly reduced growth and migration properties, whereas its overexpression demonstrated the opposite effect. Immunoprecipitation and Bimolecular fluorescence complementation (BiFC) assay confirmed the previous photo array finding of the interaction between eIF2S2 and SMAD4 to understand the tumorigenic mechanism of eIF2S2. The results indicated that the N-terminus of eIF2S2 interacts with the MH-1 domain of SMAD4. The interaction effect between eIF2S2 and SMAD4 was further evaluated. The knockdown of eIF2S2 increased SMAD4 expression in cervical cancer cells without changing SMAD4 mRNA expression, whereas transient eIF2S2 overexpression reduced SMAD4 expression. This indicates the possibility of post-translational regulation of SMAD4 expression by eIF2S2. Additionally, eIF2S2 overexpression was confirmed to weaken the expression and/or promoter activity of p15 and p27, which are SMAD4-regulated antiproliferative proteins, by reducing SMAD4 levels. Therefore, our study indicated the pro-tumorigenic role of eIF2S2, which diminishes both SMAD4 expression and function as a transcriptional factor in cervical carcinogenesis.

Maintenance of cellular ATP level by caloric restriction correlates chronological survival of budding yeast.

The free radical theory of aging emphasizes cumulative oxidative damage in the genome and intracellular proteins due to reactive oxygen species (ROS), which is a major cause for aging. Caloric restriction (CR) has been known as a representative treatment that prevents aging; however, its mechanism of action remains elusive. Here, we show that CR extends the chronological lifespan (CLS) of budding yeast by maintaining cellular energy levels. CR reduced the generation of total ROS and mitochondrial superoxide; however, CR did not reduce the oxidative damage in proteins and DNA. Subsequently, calorie-restricted yeast had higher mitochondrial membrane potential (MMP), and it sustained consistent ATP levels during the process of chronological aging. Our results suggest that CR extends the survival of the chronologically aged cells by improving the efficiency of energy metabolism for the maintenance of the ATP level rather than reducing the global oxidative damage of proteins and DNA.

Macrophage-stimulating protein attenuates gentamicin-induced inflammation and apoptosis in human renal proximal tubular epithelial cells.

The present study aimed to investigate whether macrophage-stimulating protein (MSP) treatment attenuates renal apoptosis and inflammation in gentamicin (GM)-induced tubule injury and its underlying molecular mechanisms. To examine changes in MSP and its receptor, recepteur d'origine nantais (RON) in GM-induced nephropathy, rats were injected with GM for 7 days. Human renal proximal tubular epithelial (HK-2) cells were incubated with GM for 24h in the presence of different concentrations of MSP and cell viability was measured by MTT assay. Apoptosis was determined by flow cytometry of cells stained with fluorescein isothiocyanate-conjugated annexin V protein and propidium iodide. Expression of Bcl-2, Bax, caspase-3, cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS), nuclear factor-kappa B (NF-κB), IκB-α, and mitogen-activated protein kinases (MAPKs) was analyzed by semiquantitative immunoblotting. MSP and RON expression was significantly greater in GM-treated rats, than in untreated controls. GM-treatment reduced HK-2 cell viability, an effect that was counteracted by MSP. Flow cytometry and DAPI staining revealed GM-induced apoptosis was prevented by MSP. GM reduced expression of anti-apoptotic protein Bcl-2 and induced expression of Bax and cleaved caspase 3; these effects and GM-induced expression of COX-2 and iNOS were also attenuated by MSP. GM caused MSP-reversible induction of phospho-ERK, phospho-JNK, and phospho-p38. GM induced NF-κB activation and degradation of IκB-α; the increase in nuclear NF-κB was blocked by inhibitors of ERK, JNK, p-38, or MSP pretreatment. These findings suggest that MSP attenuates GM-induced inflammation and apoptosis by inhibition of the MAPKs/NF-κB signaling pathways.

Association of age and BP variability with long-term mortality in hemodialysis patients.

BACKGROUND/AIMS: Blood pressure (BP) variability is known as a poor prognostic factor for cardiovascular outcomes. This study assessed the prognostic significance of BP variability in association with increasing age in hemodialysis patients. METHODS: We retrospectively analyzed 2,174 patients on hemodialysis from March 2005 to December 2012. The impact of intradialytic and interdialytic BP variability on all-cause mortality according to age groups was analyzed. RESULTS: Kaplan-Meier survival curves for 5-year cumulative mortality showed higher mortality in patients with higher intradialytic systolic and diastolic BP variability as well as interdialytic systolic and diastolic BP variability (log-rank p=0.006, <0.001, 0.018 and < 0.001) in patients aged <55 years, but not in older age groups. Cox proportional analysis revealed that 5-year mortality was associated with intradialytic diastolic BP variability in patients aged <55 years (HR, 2.03 CI, 1.24-3.32). CONCLUSION: The overall mortality was associated with BP variability in patients aged <55 years, but not in older ages. This result suggests that younger hemodialysis patients with BP variability require further medical attention and intervention to reduce BP variability.

Prevalence and associations for abnormal bleeding times in patients with renal insufficiency.

Platelet dysfunction and associated hemorrhagic complications are often encountered in patients with chronic kidney disease. This study aimed to evaluate the prevalence and associations for abnormal bleeding time (BT) in patients with renal dysfunction. Hemoglobin, hematocrit, platelet, blood urea nitrogen, creatinine, and parathyroid hormone levels were determined in 1716 patients (55.18 ± 17.19 years, men 50.8%). For these patients, BTs were estimated using a platelet function analyzer-100. Glomerular filtration rates (GFRs) were estimated using the Chronic Kidney Disease Epidemiology Collaboration equation. The study population was divided into six groups according to the estimated GFR (eGRF): group I, eGFR ≥ 90 ml/min/1.73 m(2); group II, 60 ≤ eGFR < 90 ml/min/1.73 m(2); group III, 30 ≤ eGFR < 60 ml/min/1.73 m(2); group IV, 15 ≤ eGFR < 30 ml/min/1.73 m(2); group V, eGFR < 15 ml/min/1.73 m(2); and group VI, undergoing regular hemodialysis. Renal insufficiency was defined as eGFR < 60 ml/min/1.73 m(2). To further investigate the role of inflammatory cytokines, nitric oxide (NO) and tumor necrosis factor alpha (TNF-α) were measured in a 327-patient subset of the total patient population (52.82 ± 18.3 years, men 60.9%). Abnormal BT occurred in 11.8% of group I, 15.3% of group II, 29.1% of group III, 37.5% of group IV, 35.0% of group V, and 32.1% of group VI. By Pearson correlation coefficient, eGFR (r = -0.089), hemoglobin (r = -0.127), platelet (r = -0.054) were correlated with BT. Multivariate analysis revealed that age [odds ratio (OR), 1.013; 95% CI, 1.004-1.022], renal insufficiency (eGFR < 60 ml/min/1.73 m(2); OR, 2.271; 95% CI, 1.672-3.083), anemia (hemoglobin < 120 g/l; OR, 1.486; 95% CI, 1.089-2.027), and thrombocytopenia (platelet < 150 × 10(9)/l; OR, 1.445; 95% CI, 1.089-1.918) were independently associated with prolonged BT. Plasma levels of NO and TNF-α were increased in patients with renal insufficiency (eGFR < 60 ml/min/1.73 m(2)). Plasma levels of NO in renal insufficiency group were higher in prolonged BT than those in normal BT. A significant positive correlation was noted between BTs and NO levels (r = 0.152, p = 0.009) but not with TNF-α levels. The prevalence of abnormal BTs was higher as eGFR declined. Old age, renal insufficiency, anemia, and thrombocytopenia were independent associations for abnormal BT.

Treatment of Dowling-Degos disease with fractional Er:YAG laser.

Dowling-Degos disease (DDD) is a rare, benign, autosomal dominant disorder characterized by reticulated pigmentation on flexural areas. Recently, a report of successful Er:YAG ablative laser treatment without any adverse effects was issued. A 49-year-old Korean woman presented with numerous small, hyperpigmented macules in a reticular pattern on her face, axillae, and inguinal folds of several years duration. Histologic examination revealed acanthosis with thin elongated rete ridges, basal branching, and widening. She was diagnosed as having DDD and was treated successfully without any adverse effects using a fractional 2,940-nm Er:YAG ablative laser (LOTUSII, Laseroptek, Sungnam, Korea). Er:YAG ablative laser treatment could be an effective treatment modality for DDD, but in Asians, who have darker skins than Caucasians, or in patients with skin lesions on the face, post-inflammatory hyperpigmentation could be problematic after treatment. Fractional Er:YAG ablative laser treatment should be viewed as an alternative therapeutic option for DDD.