Tumoricidal Activity of Simvastatin in Synergy with RhoA Inactivation in Antimigration of Clear Cell Renal Cell Carcinoma Cells.

Among kidney cancers, clear cell renal cell carcinoma (ccRCC) has the highest incidence rate in adults. The survival rate of patients diagnosed as having metastatic ccRCC drastically declines even with intensive treatment. We examined the efficacy of simvastatin, a lipid-lowering drug with reduced mevalonate synthesis, in ccRCC treatment. Simvastatin was found to reduce cell viability and increase autophagy induction and apoptosis. In addition, it reduced cell metastasis and lipid accumulation, the target proteins of which can be reversed through mevalonate supplementation. Moreover, simvastatin suppressed cholesterol synthesis and protein prenylation that is essential for RhoA activation. Simvastatin might also reduce cancer metastasis by suppressing the RhoA pathway. A gene set enrichment analysis (GSEA) of the human ccRCC GSE53757 data set revealed that the RhoA and lipogenesis pathways are activated. In simvastatin-treated ccRCC cells, although RhoA was upregulated, it was mainly restrained in the cytosolic fraction and concomitantly reduced Rho-associated protein kinase activity. RhoA upregulation might be a negative feedback effect owing to the loss of RhoA activity caused by simvastatin, which can be restored by mevalonate. RhoA inactivation by simvastatin was correlated with decreased cell metastasis in the transwell assay, which was mimicked in dominantly negative RhoA-overexpressing cells. Thus, owing to the increased RhoA activation and cell metastasis in the human ccRCC dataset analysis, simvastatin-mediated Rho inactivation might serve as a therapeutic target for ccRCC patients. Altogether, simvastatin suppressed the cell viability and metastasis of ccRCC cells; thus, it is a potentially effective ccRCC adjunct therapy after clinical validation for ccRCC treatment.

Sigesbeckia orientalis Extract Ameliorates the Experimental Diabetic Nephropathy by Downregulating the Inflammatory and Oxidative Stress Signaling Pathways.

Diabetes in children and its complications are on the rise globally, which is accompanied by increasing in diabetes-related complications. Oxidative stress and inflammation induced by elevated blood sugar in diabetic patients are considered risk factors associated with the development of diabetes complications, including chronic kidney disease and its later development to end-stage renal disease. Microvascular changes within the kidneys of DM patients often lead to chronic kidney disease, which aggravates the illness. Sigesbeckia orientalis extract (SOE), reported to have strong antioxidative and excellent anti-inflammatory activities, is used in the modern practice of traditional Chinese medicine. Kidneys from three groups of control mice (CTR), mice with streptozotocin (STZ)-induced diabetes (DM), and mice with STZ-induced DM treated with SOE (DMRx) were excised for morphological analyses and immunohistochemical assessments. Only mice in the DM group exhibited significantly lower body weight, but higher blood sugar was present. The results revealed more obvious renal injury in the DM group than in the other groups, which appeared as greater glomerular damage and tubular injury, sores, and plenty of connective tissues within the mesangium. Not only did the DM group have a higher level of cytokine, tumor necrosis factor, and the oxidative stress marker, 8-hydroxyguanosine expression, but also factors of the nuclear factor pathway and biomarkers of microvascular status had changed. Disturbances to the kidneys in DMRx mice were attenuated compared to the DM group. We concluded that SOE is an effective medicine, with antioxidative and anti-inflammatory abilities, to protect against or attenuate diabetic nephropathy from inflammatory disturbances by oxidative stress and to cure vessel damage in a hyperglycemic situation.

L-Carnitine reduces reactive oxygen species/endoplasmic reticulum stress and maintains mitochondrial function during autophagy-mediated cell apoptosis in perfluorooctanesulfonate-treated renal tubular cells.

We previously reported that perfluorooctanesulfonate (PFOS) causes autophagy-induced apoptosis in renal tubular cells (RTCs) through a mechanism dependent on reactive oxygen species (ROS)/extracellular signal-regulated kinase. This study extended our findings and determined the therapeutic potency of L-Carnitine in PFOS-treated RTCs. L-Carnitine (10 mM) reversed the effects of PFOS (100 µM) on autophagy induction and impaired autophagy flux. Furthermore, it downregulated the protein level of p47Phox, which is partly related to PFOS-induced increased cytosolic ROS in RTCs. Moreover, L-Carnitine reduced ROS production in mitochondria and restored PFOS-impeded mitochondrial function, leading to sustained normal adenosine triphosphate synthesis and oxygen consumption and reduced proton leakage in a Seahorse XF stress test. The increased inositol-requiring enzyme 1α expression by PFOS, which indicated endoplasmic reticulum (ER) stress activation, was associated with PFOS-mediated autophagy activation that could be attenuated through 4-phenylbutyrate (5 mM, an ER stress inhibitor) and L-Carnitine pretreatment. Therefore, by reducing the level of IRE1α, L-Carnitine reduced the levels of Beclin and LC3BII, consequently reducing the level of apoptotic biomarkers including Bax and cleaving PARP and caspase 3. Collectively, these results indicate that through the elimination of oxidative stress, extracellular signal-regulated kinase activation, and ER stress, L-Carnitine reduced cell autophagy/apoptosis and concomitantly increased cell viability in RTCs. This study clarified the potential mechanism of PFOS-mediated RTC apoptosis and provided a new strategy for using L-Carnitine to prevent and treat PFOS-induced RTC apoptosis.

Glossogyne tenuifolia Extract Increases Nitric Oxide Production in Human Umbilical Vein Endothelial Cells.

The vascular nitric oxide (NO) system has a protective effect in atherosclerosis. NO is generated from the conversion of L-arginine to L-citrulline by the enzymatic action of endothelial NO synthase (eNOS). Compounds with the effect of enhancing eNOS expression are considered to be candidates for the prevention of atherosclerosis. In this study, extracts from the aerial, root, and whole plant of Glossogyne tenuifolia (GT) were obtained with ethanol, n-hexane, ethyl acetate (EA), and methanol extraction, respectively. The effects of these GT extracts on the synthesis of NO and the expression of eNOS in human umbilical vein endothelial cells (HUVECs) were investigated. NO production was determined as nitrite by colorimetry, following the Griess reaction. The treatment of HUVECs with EA extract from the root of GT and n-hexane, methanol, and ethanol extract from the aerial, root, and whole plant of GT increased NO production in a dose-dependent manner. When at a dose of 160 µg/mL, NO production increased from 0.9 to 18.4-fold. Among these extracts, the methanol extract from the root of GT (R/M GTE) exhibited the most potent effect on NO production (increased by 18.4-fold). Furthermore, using Western blot and RT-PCR analysis, treatment of HUVECs with the R/M GTE increased both eNOS protein and mRNA expression. In addition, Western blot analysis revealed that the R/M GTE increased eNOS phosphorylation at serine1177 as early as 15 min after treatment. The chemical composition for the main ingredients was also performed by HPLC analysis. In conclusion, the present study demonstrated that GT extracts increased NO production in HUVECs and that the R/M GTE increased NO production via increasing eNOS expression and activation by phosphorylation of eNOS at serine1177.

Perfluorooctane sulfonate induces autophagy-associated apoptosis through oxidative stress and the activation of extracellular signal-regulated kinases in renal tubular cells.

Perfluorooctane sulfonate (PFOS) is among the most abundant organic pollutants and is widely distributed in the environment, wildlife, and humans. Its toxic effects and biological hazards are associated with its long elimination half-life in humans. However, how it affects renal tubular cells (RTCs) remains unclear. In this study, PFOS was observed to mediate the increase in reactive oxygen species (ROS) generation, followed by the activation of the extracellular-signal-regulated kinase 1/2 (ERK1/2) pathway, which induced autophagy in RTCs. Although PFOS treatment induced autophagy after 6 h, prolonged treatment (24 h) reduced the autophagic flux by increasing lysosomal membrane permeability (LMP), leading to increased p62 protein accumulation and subsequent apoptosis. The increase in LMP was visualized through increased green fluorescence with acridine orange staining, and this was attenuated by 3-methyladenine, an autophagy inhibitor. N-acetyl cysteine and an inhibitor of the mitogen-activated protein kinase kinases (U0126) attenuated autophagy and apoptosis. Taken together, these results indicate that ROS activation and ROS-mediated phosphorylated ERK1/2 activation are essential to activate autophagy, resulting in the apoptosis of PFOS-treated RTCs. Our findings provide insight into the mechanism of PFOS-mediated renal toxicity.

Antrodia cinnamomea Extraction Waste Supplementation Promotes Thermal Stress Tolerance and Tissue Regeneration Ability of Zebrafish.

Antrodia cinnamomea (AC) has been shown to have anti-inflammatory, anti-tumor, and immunomodulation activities. It is estimated that hundreds of metric tons of AC extraction waste (ACEW) are produced per year in Taiwan. This study aims to assess the feasibility of applying ACEW as feed supplement in the aquaculture industry. ACEW significantly inhibited the growth of microorganisms in the water tank, by around 39.4% reduction on the fifth day with feed supplemented of 10% ACEW. The feed conversion efficiency of zebrafish with 10% ACEW supplementation for 30 days was 1.22-fold compared to that of the control. ACEW dramatically improved the tolerances of zebrafish under the heat and cold stresses. When at water temperature extremes of 38 °C or 11 °C, compared to the 100% mortality rate in the control group, the 10% ACEW diet group still had 91.7% and 83.3% survival rates, respectively. In a caudal fin amputation test, the fin recovery of zebrafish was increased from 68.4% to 93% with 10% ACEW diet after 3-week regeneration. ACEW effectively down-regulated the gene expression of TNF-α, IL-1ß, IL-6, and IL-10, and up-regulated the gene expression of IL-4/13A. Additionally, the supplement of ACEW in the feed can maintain and prevent the fish's body weight from dropping too much under enteritis. Taken together, ACEW has beneficial potential in aquaculture.

Reversal of bleomycin-induced rat pulmonary fibrosis by a xenograft of human umbilical mesenchymal stem cells from Wharton's jelly.

Pulmonary fibrosis (PF) is a progressive and irreversible condition with various causes, and no effective treatment has been found to rescue fibrotic lungs. Successful recovery from PF requires inhibiting inflammation, promoting collagen degradation and stimulating alveolar regeneration. Human umbilical mesenchymal stem cells (HUMSCs) not only regulate immune responses but also synthesize and release hyaluronan to improve lung regeneration. This study investigated the feasibility of HUMSC engraftment into rats with bleomycin (BLM)-induced PF to explore HUMSC therapeutic effects/outcomes. Methods: A unique BLM-induced left-lung-dominated PF animal model was established. Rats were transplanted with low-dose (5×106) or high-dose (2.5×107) HUMSCs on Day 21 after BLM injection. Combinations in co-culture of pulmonary macrophages, fibroblasts, HUMSCs treated with BLM and the same conditions on alveolar epithelia versus HUMSCs were evaluated. Results: Rats with high-dose HUMSC engraftment displayed significant recovery, including improved blood oxygen saturation levels and respiratory rates. High-dose HUMSC transplantation reversed alveolar injury, reduced cell infiltration and ameliorated collagen deposition. One month posttransplantation, HUMSCs in the rats' lungs remained viable and secreted cytokines without differentiating into alveolar or vascular epithelial cells. Moreover, HUMSCs decreased epithelial-mesenchymal transition in pulmonary inflammation, enhanced macrophage matrix-metallopeptidase-9 (MMP-9) expression for collagen degradation, and promoted toll-like receptor-4 (TLR-4) expression in the lung for alveolar regeneration. In coculture studies, HUMSCs elevated the MMP-9 level in pulmonary macrophages, released hyaluronan into the medium and stimulated the TLR-4 quantity in the alveolar epithelium. Principal Conclusions: Transplanted HUMSCs exhibit long-term viability in rat lungs and can effectively reverse rat PF.

Xenografting of human umbilical mesenchymal stem cells from Wharton's jelly ameliorates mouse spinocerebellar ataxia type 1.

BACKGROUND: Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disorder caused by the expansion of CAG repeats in ATXN1 gene resulting in an expansion of polyglutamine repeats in the ATXN1 protein. Unfortunately, there has yet been any effective treatment so far for SCA1. This study investigated the feasibility of transplanting human umbilical mesenchymal stem cells (HUMSCs) into transgenic SCA1 mice containing an expanded uninterrupted allele with 82 repeats in the ATXN1-coding region. METHODS: 106 human umbilical mesenchymal stem cells were transplanted into the cerebella at 1 month of age. RESULTS: HUMSCs displayed significant ameliorating effects in SCA1 mice in terms of motor behaviors in balance beam test and open field test as compared with the untransplanted SCA1 mice. HUMSCs transplantation effectively reduced the cerebellar atrophy, salvaged Purkinje cell death, and alleviated molecular layer shrinkage. Electrophysiological studies showed higher amplitudes of compound motor action potentials as indicated by increasing neuronal-muscular response strength to stimuli after stem cell transplantation. At 5 months after transplantation, HUMSCs scattering in the mice cerebella remained viable and secreted cytokines without differentiating into neuronal or glia cells. CONCLUSIONS: Our findings provide hope for a new therapeutic direction for the treatment of SCA1.

Xenograft of Human Umbilical Mesenchymal Stem Cells from Wharton's Jelly Differentiating into Osteocytes and Reducing Osteoclast Activity Reverses Osteoporosis in Ovariectomized Rats.

We examined the effects of human umbilical cord-derived mesenchymal stem cells (HUMSCs) in Wharton's jelly on ovariectomy (OVX)-induced osteoporosis by using in vitro and in vivo experiments. Two months after OVX, the rats gained weight and had a decreased serum estradiol level . Both micro-computed tomography (micro-CT) and histochemical analyses revealed a marked decrease in the bone volume (BV) and collagen content within the head, neck, and distal condyle of the femur, indicating that the osteoporosis animal model was successfully established 2 mo after bilateral OVX. Subsequently, 2.5 × 106 HUMSCs were injected into the bone marrow cavity of the left femurs 2 mo after OVX. The rats were divided into the following groups: normal + phosphate-buffered saline (PBS), normal + HUMSCs, OVX + PBS, and OVX + HUMSCs. Two months after transplantation, both micro-CT imaging and histochemical staining revealed that the normal + HUMSCs group had higher BV and collagen content in the epiphysis and metaphysis than did the normal + PBS group. In the OVX + HUMSCs group, a substantial increase in the rod-shaped trabecular bone and the abundant accumulation of collagen were observed around the site of HUMSC transplantation. Plenty of transplanted HUMSCs remained viable and differentiated into osteoblasts. In addition, HUMSC transplantation reduced the number of osteoclasts. Compared with HUMSCs cultured alone, HUMSCs cocultured with osteoblasts showed that the percentage of cells differentiating into osteoblasts significantly increased. Furthermore, osteoclasts cocultured with HUMSCs had significantly decreased cellular activity and differentiation capability. HUMSC transplantation into the distal femur of OVX rats could locally stimulate osteocalcin synthesis, increase the trabecular bone, and inhibit osteoclast activity.

Xenograft of human umbilical mesenchymal stem cells from Wharton's jelly as a potential therapy for rat pilocarpine-induced epilepsy.

We evaluated the effects of intra-hippocampal transplantation of human umbilical mesenchymal stem cells (HUMSCs) on pilocarpine-treated rats. Sprague-Dawley rats were divided into the following three groups: (1) a normal group of rats receiving only PBS, (2) a status epilepticus (SE) group of rats with pilocarpine-induced SE and PBS injected into the hippocampi, and (3) a SE+HUMSC group of SE rats with HUMSC transplantation. Spontaneous recurrent motor seizures (SRMS) were monitored using simultaneous video and electroencephalographic recordings at two to four weeks after SE induction. The results showed that the number of SRMS within two to four weeks after SE was significantly decreased in SE+HUMSCs rats compared with SE rats. All of the rats were sacrificed on Day 29 after SE. Hippocampal morphology and volume were evaluated using Nissl staining and magnetic resonance imaging. The results showed that the volume of the dorsal hippocampus was smaller in SE rats compared with normal and SE+HUMSCs rats. The pyramidal neuron loss in CA1 and CA3 regions was more severe in the SE rats than in normal and SE+HUMSCs rats. No significant differences were found in the hippocampal neuronal loss or in the number of dentate GABAergic neurons between normal and SE+HUMSCs rats. Compared with the SE rats, the SE+HUMSCs rats exhibited a suppression of astrocyte activity and aberrant mossy fiber sprouting. Implanted HUMSCs survived in the hippocampus and released cytokines, including FGF-6, amphiregulin, glucocorticoid-induced tumor necrosis factors receptor (GITR), MIP-3ß, and osteoprotegerin. In an in vitro study, exposure of cortical neurons to glutamate showed a significant decrease in cell viability, which was preventable by co-culturing with HUMSCs. Above all, the expression of human osteoprotegerin and amphiregulin were significantly increased in the media of the co-culture of neurons and HUMSCs. Our results demonstrate the therapeutic benefits of HUMSC transplantation for the development of epilepsy, which are likely due to the ability of the cells to produce neuroprotective and anti-inflammatory cytokines. Thus, HUMSC transplantation may be an effective therapy in the future.

The Therapeutic Potential of Human Umbilical Mesenchymal Stem Cells From Wharton's Jelly in the Treatment of Rat Peritoneal Dialysis-Induced Fibrosis.

A major complication in continuous, ambulatory peritoneal dialysis in patients with end-stage renal disease who are undergoing long-term peritoneal dialysis (PD) is peritoneal fibrosis, which can result in peritoneal structural changes and functional ultrafiltration failure. Human umbilical mesenchymal stem cells (HUMSCs) in Wharton's jelly possess stem cell properties and are easily obtained and processed. This study focuses on the effects of HUMSCs on peritoneal fibrosis in in vitro and in vivo experiments. After 24-hour treatment with mixture of Dulbecco's modified Eagle's medium and PD solution at a 1:3 ratio, primary human peritoneal mesothelial cells became susceptible to PD-induced cell death. Such cytotoxic effects were prevented by coculturing with primary HUMSCs. In a rat model, intraperitoneal injections of 20 mM methylglyoxal (MGO) in PD solution for 3 weeks (the PD/MGO 3W group) markedly induced abdominal cocoon formation, peritoneal thickening, and collagen accumulation. Immunohistochemical analyses indicated neoangiogenesis and significant increase in the numbers of ED-1- and α-smooth muscle actin (α-SMA)-positive cells in the thickened peritoneum in the PD/MGO 3W group, suggesting that PD/MGO induced an inflammatory response. Furthermore, PD/MGO treatment for 3 weeks caused functional impairments in the peritoneal membrane. However, in comparison with the PD/MGO group, intraperitoneal administration of HUMSCs into the rats significantly ameliorated the PD/MGO-induced abdominal cocoon formation, peritoneal fibrosis, inflammation, neoangiogenesis, and ultrafiltration failure. After 3 weeks of transplantation, surviving HUMSCs were found in the peritoneum in the HUMSC-grafted rats. Thus, xenografts of HUMSCs might provide a potential therapeutic strategy in the prevention of peritoneal fibrosis. Significance: This study demonstrated that direct intraperitoneal transplantation of human umbilical mesenchymal stem cells into the rat effectively prevented peritoneal dialysis/methylglyoxal-induced abdominal cocoon formation, ultrafiltration failure, and peritoneal membrane alterations such as peritoneal thickening, fibrosis, and inflammation. These findings provide a basis for a novel approach for therapeutic benefits in the treatment of encapsulating peritoneal sclerosis.

A high-efficiency induction of dopaminergic cells from human umbilical mesenchymal stem cells for the treatment of hemiparkinsonian rats.

The success rate in previous attempts at transforming human umbilical mesenchymal stem cells (HUMSCs) isolated from Wharton's jelly of the umbilical cord into dopaminergic cells was a mere 12.7%. The present study was therefore initiated to establish a more effective procedure for better yield of dopaminergic cells in such transformation for more effective HUMSC-based therapy for parkinsonism. To examine, in vitro, the effects of enhanced Nurr1 expression in HUMSCs on their differentiation, cells were processed through the three-stage differentiation protocol. The capacity of such cells to synthesize and release dopamine was measured by HPLC. The therapeutic effects of Nurr1-overexppressed HUMSCs were examined in 6-hydroxydopamine-lesioned rats by quantification of rotations in response to amphetamine. Enhanced Nurr1 expression in HUMSCs promoted the transformation into dopaminergic cells in vitro through stepwise culturing in sonic hedgehog, fibroblast growth factor-8, and neuron-conditioned medium. The success rate was about 71%, as determined by immunostaining for tyrosine hydroxylase and around 94 nM dopamine synthesis (intracellular and released into the culture medium), as measured by HPLC. Additionally, transplantation of such cells into the striatum of hemiparkinsonian rats resulted in improvement of their behavioral deficits, as indicated by amphetamine-evoked rotation scores. Viability of the transplanted cells lasted for at least 3 months as verified by positive staining for tyrosine hydroxylase. Nurr1, FGF8, Shh, and NCM can synergistically enhance the differentiation of HUMSCs into dopaminergic cells and may pave the way for HUMSC-based treatments for Parkinson's disease.

Valproic acid suppresses lipopolysaccharide-induced cyclooxygenase-2 expression via MKP-1 in murine brain microvascular endothelial cells.

Inflammation and vascular perturbations are increasingly implicated in the pathogenesis of neurodegenerative diseases. Prevailing evidence suggests that valproic acid (VPA), an antiepileptic and mood stabilizer, exhibits not only neuro-protective effects, but also anti-inflammatory effects in neurodegenerative diseases. However, the underlying mechanism contributing to VPA's suppression of inflammatory responses remains unclear. In this study, we explored the inhibitory action of VPA on cyclooxygenase (COX)-2 expression in bEnd.3 mouse brain microvascular endothelial cells exposed to lipopolysaccharide (LPS), a pro-inflammatory stimulus. The LPS-induced increases in COX-2 protein level and COX-2 promoter-luciferase activity were significantly suppressed by VPA. VPA inhibited p38MAPK and JNK phosphorylation in LPS-stimulated bEnd.3 cells. Treatment of cells with a p38MAPK inhibitor (p38MAPK inhibitor III) or a JNK signaling inhibitor (JNK inhibitor II) significantly inhibited LPS-induced COX-2 expression. VPA inhibited LPS-induced NF-κB subunit p65 phosphorylation and κB-luciferase activity. LPS-increased p65 and C/EBPß binding to the COX-2 promoter region was attenuated in the presence of VPA. In addition, VPA suppression of p38MAPK, JNK and p65 phosphorylation, and subsequent COX-2 expression was restored in cells transfected with mitogen-activated protein kinase phosphatase-1 (MKP-1) dominant negative (DN) mutant. VPA also caused increases in MKP-1 acetylation and MKP-1 phosphatase activity in bEnd.3 cells. In conclusion, VPA may cause MKP-1 activation to dephosphorylate p38MAPK and JNK, leading to decrease in p65 and C/EBPß binding to the COX-2 promoter region and COX-2 down-regulation in LPS-stimulated bEnd.3 cells. The present study therefore supports the therapeutic value of VPA in alleviating brain inflammatory processes.

Effects of alanyl-glutamine dipeptide on the expression of colon-inflammatory mediators during the recovery phase of colitis induced by dextran sulfate sodium.

PURPOSE: Glutamine (Gln) is a nutrient with immunomodulatory effects in metabolic stressed conditions. This study investigated the effects of Gln on colonic-inflammatory-mediator expression and mucosal repair in mice with dextran sulfate sodium (DSS)-induced colitis. METHODS: C57BL/6 mice received distilled water containing 3 % DSS for 5 d to induce colitis. One of the DSS-treated groups was intraperitoneally injected with an alanyl (Ala)-Gln solution 3 days before (G-DSS) while the other group was administered Ala-Gln 3 days after colitis (DSS-G) was induced. The Ala-Gln solution provided 0.5 g Gln/kg/d. The saline-DSS group (S-DSS) received an identical amount of saline before and after colitis was induced to serve as a positive control. RESULTS: The S-DSS group had a shorter colon length, higher plasma haptoglobin level, and more-severe colon inflammation. Also, the toll-like receptor (TLR)4 level, nuclear factor (NF)-κB activation, and inflammatory cytokine gene expression in the colon were higher than those of the normal control group. Gln administration either before or after colitis suppressed TLR4 protein levels, decreased plasma haptoglobin, and reduced colon inflammation. Histological inflammatory scores were also lowered. Compared to the post-colitis Gln group, preventive use of Gln had higher colon length, expressions of mucin 2, trefoil factor 3, and heat shock protein 72 genes were also upregulated in the colon. CONCLUSIONS: These results suggest that Gln administered either before or after the colitis mitigated inflammation of colitis that was not observed in group without Gln injection. Prophylactic treatment with Gln had more-beneficial effects on reducing inflammatory markers and enhancing the recovery of mucosa in DSS-induced colitis.

Glutamine modulates sepsis-induced changes to intestinal intraepithelial γδT lymphocyte expression in mice.

This study investigated the effect of glutamine (GLN) on intestinal intraepithelial lymphocyte (IEL) γδT-cell cytokines and immune regulatory factor gene expressions in a mouse model of polymicrobial sepsis. Mice were randomly assigned to a normal group, a sepsis with saline (SS) group, or a sepsis with GLN (SG) group. All mice were fed a chow diet. Sepsis was induced by cecal ligation and puncture (CLP). The SS group was injected with saline, and the SG group was given 0.75 g GLN/kg body weight once via a tail vein 1 h after CLP. Septic mice were killed 12 h after CLP, and IEL γδT cells of the animals were isolated for further analysis. Results showed that compared with normal mice, sepsis resulted in lower IEL γδT-cell percentage and higher messenger RNA expressions of interferon γ, tumor necrosis factor α, interleukin 4 (IL-4), IL-13, IL-17, retinoid acid receptor-related orphan receptor γt, and complement 5a receptor by IEL γδT cells. These immunomodulatory mediator genes exhibited decreases, whereas IL-7 receptor expression increased in IEL γδT cells in septic mice with GLN administration. Annexin V/7-amino-actinomycin D stain revealed significantly lower rates of apoptosis, and IEL γδT-cell percentage was higher in the SG group. The histological findings also showed that damage to intestinal epithelial cells was less severe in the SG group. These results indicated that a single dose of GLN administered as treatment after the initiation of sepsis prevented apoptosis of IEL γδT cells and downregulated γδT cell-expressed inflammatory mediators that may consequently ameliorate the severity of sepsis-induced intestinal epithelial injury.

Postnatal development of microcyst in the anteroventral cochlear nucleus of the Mongolian gerbil: a light- and electron microscopic study.

We investigated the postnatal formation and origin of the microcyst, which are not fully elucidated at present, in the cochlear nucleus of gerbils. Sixty-six Mongolian gerbils were investigated at the light microscope level, and 35 of them were observed at the electron microscopic level. Foamy structures were evidently found at 2 days of age and remained unchanged through 4-8 days. The first small vacuole, presumably the former microcyst, appeared at 8 days. Myelin sheath bundles first appeared at 13 days. Electron-dense bodies were frequently found in the junction of the superficial layer and the deep layer at 2 days. The medium-sized vacuole was found in close association with the spherical bushy cells in the anteroventral cochlear nucleus (AVCN) as early as 5 weeks. Various large and small vacuoles were presumably coalesced to form a large vacuole at 3 and 6 months. Membranous structures and red blood cells were in the budding-like vacuoles at 6 months. In addition to membranous structures, the microcyst contained distorted mitochondria and parts of myelin sheaths. The vacuole was interposed between spherical bushy cells at age of 10 months. Small vacuoles were mainly located in the flame-shaped neurons at 14 months. An internal detachment and an external protrusion of the myelin sheath into the adjacent microcyst were found. Thus, this study suggests the first appearance of microcysts at 8 days. Also, the microcyst and the blood vessel may exchange their contents through a leakage in the anteroventral cochlear nucleus.

Human umbilical mesenchymal stem cells promote recovery after ischemic stroke.

BACKGROUND AND PURPOSE: Stroke is a cerebrovascular defect that leads to many adverse neurological complications. Current pharmacological treatments for stroke remain unclear in their effectiveness, whereas stem cell transplantation shows considerable promise. Previously, we have shown that human umbilical mesenchymal stem cells (HUMSCs) can differentiate into neurons in neuronal-conditioned medium. Here we evaluate the therapeutic potential of HUMSC transplantation for ischemic stroke in rats. METHODS: Focal cerebral ischemia was produced by middle cerebral artery occlusion and reperfusion. The HUMSCs treated with neuronal-conditioned medium or not treated were transplanted into the ischemic cortex 24 hours after surgery. RESULTS: Histology and MRI revealed that rats implanted with HUMSCs treated with neuronal-conditioned medium or not treated exhibited a trend toward less infarct volume and significantly less atrophy compared with the control group, which received no HUMSCs. Moreover, rats receiving HUMSCs showed significant improvements in motor function, greater metabolic activity of cortical neurons, and better revascularization in the infarct cortex. Implanted HUMSCs, treated or not treated, survived in the infarct cortex for at least 36 days and released neuroprotective and growth-associated cytokines, including brain-derived neurotrophic factor, platelet-derived growth factor-AA, basic fibroblast growth factor, angiopoietin-2, CXCL-16, neutrophil-activating protein-2, and vascular endothelial growth factor receptor-3. CONCLUSIONS: Our results demonstrate the therapeutic benefits of HUMSC transplantation for ischemic stroke, likely due to the ability of the cells to produce growth-promoting factors. Thus, HUMSC transplantation may be an effective therapy in the future.

The therapeutic potential of human umbilical mesenchymal stem cells from Wharton's jelly in the treatment of rat liver fibrosis.

We investigated the effect of human umbilical mesenchymal stem cells (HUMSCs) from Wharton's jelly on carbon tetrachloride (CCl4)-induced liver fibrosis in rats. Rats were treated with CCl4 for 4 weeks, and this was followed by a direct injection of HUMSCs into their livers. After 4 more weeks of CCl4 treatment (8 weeks in all), rats with HUMSC transplants [CCl4 (8W)+HUMSC liver] exhibited a significant reduction in liver fibrosis, as evidenced by Sirius red staining and a collagen content assay, in comparison with rats treated with CCl4 for 8 weeks without HUMSC transplants [CCl4 (8W)]. Moreover, rats in the CCl4 (8W)+HUMSC (liver) group had significantly lower levels of serum glutamic oxaloacetic transaminase, glutamic pyruvate transaminase, alpha-smooth muscle actin, and transforming growth factor-beta1 in the liver, whereas the expression of hepatic mesenchymal epithelial transition factor-phosphorylated type (Met-P) and hepatocyte growth factor was up-regulated, in comparison with the CCl4 (8W) group. Notably, engrafted HUMSCs scattered mostly in the hepatic connective tissue but did not differentiate into hepatocytes expressing human albumin or alpha-fetoprotein. Instead, these engrafted, undifferentiated HUMSCs secreted a variety of bioactive cytokines that may restore liver function and promote regeneration. Human cytokine assay revealed that the amounts of human cutaneous T cell-attracting chemokine, leukemia inhibitory factor, and prolactin were substantially greater in the livers of the CCl4 (8W)+HUMSC (liver) group, with considerably reduced hepatic inflammation manifested by a micro positron emission tomography scan. Our findings suggest that xenogeneic transplantation of HUMSCs is a novel approach for treating liver fibrosis and may be a promising therapeutic intervention in the future.

Identification of genetic networks during mesenchymal stem cell transformation into neurons.

The aim of this experiment is to identify related genes for human umbilical mesenchymal stem cells transformation into nervous cells. After the human umbilical mesenchymal stem cells were treated with neuronal conditioned medium (NCM) for 9 days, the gene expression groups are compared to those only treated with DMEM. The related genes for cell cycles, the human umbilical mesenchymal stem cells treated with DMEM increases the amount of cells that remain in the G2/M phase and S phase, including CAV1, EBF, NRG1, CDH13, MLH1. After treatment, the human umbilical cord mesenchymal stem cells with NCM for 9 days, gene expression related to the G0/G1 phase are also increased, including MYC, CSF3, PETN. Gene expressions related to neural regeneration and neural stem cells also increase significantly, such as CXCL1, BMP2, NRCAM, FGF2, SPG7. This study thereby provides a foundation for a more detailed understanding of HUMSCs neuronal differentiation.

Effects of streptozotocin-induced diabetes on taste buds in rat vallate papillae.

Some studies have documented taste changes in patients with diabetes mellitus (DM). In order to understand the relationships between taste disorders caused by DM and the innervation and morphologic changes in the taste buds, we studied the vallate papillae and their taste buds in rats with DM. DM was induced in these rats with streptozotocin (STZ), which causes the death of beta cells of the pancreas. The rats were sacrificed and the vallate papillae were dissected for morphometric and quantitative immunohistochemical analyses. The innervations of the vallate papillae and taste buds in diabetic and control rats were detected using immunohistochemistry employing antibodies directed against protein gene product 9.5 (PGP 9.5) and calcitonin gene-related peptide (CGRP). The results showed that PGP 9.5- and CGRP-immunoreactive nerve fibers in the trench wall of diabetic vallate papillae, as well as taste cells in the taste buds, gradually decreased both intragemmally and intergemmally. The morphometry revealed no significant difference in papilla size between the control and diabetic groups, but there were fewer taste buds per papilla (per animal). The quantification of innervation in taste buds of the diabetic rats supported the visual assessment of immunohistochemical labeling, that the innervation of taste cells was significantly reduced in diabetic animals. These findings suggest that taste impairment in diabetic subjects may be caused by neuropathy defects and/or morphological changes in the taste buds.