Improvement Effects of Myelophil on Symptoms of Chronic Fatigue Syndrome in a Reserpine-Induced Mouse Model.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is associated with various symptoms, such as depression, pain, and fatigue. To date, the pathological mechanisms and therapeutics remain uncertain. The purpose of this study was to investigate the effect of myelophil (MYP), composed of Astragali Radix and Salviaemiltiorrhizae Radix, on depression, pain, and fatigue behaviors and its underlying mechanisms. Reserpine (2 mg/kg for 10 days, intraperitoneally) induced depression, pain, and fatigue behaviors in mice. MYP treatment (100 mg/kg for 10 days, intragastrically) significantly improved depression behaviors, mechanical and thermal hypersensitivity, and fatigue behavior. MYP treatment regulated the expression of c-Fos, 5-HT1A/B receptors, and transforming growth factor ß (TGF-ß) in the brain, especially in the motor cortex, hippocampus, and nucleus of the solitary tract. MYP treatment decreased ionized calcium binding adapter molecule 1 (Iba1) expression in the hippocampus and increased tyrosine hydroxylase (TH) expression and the levels of dopamine and serotonin in the striatum. MYP treatment altered inflammatory and anti-oxidative-related mRNA expression in the spleen and liver. In conclusion, MYP was effective in recovering major symptoms of ME/CFS and was associated with the regulation of dopaminergic and serotonergic pathways and TGF-ß expression in the brain, as well as anti-inflammatory and anti-oxidant mechanisms in internal organs.

Neuroprotective effect of combined use of nicotine and celecoxib by inhibiting neuroinflammation in ischemic rats.

INTRODUCTION: The contribution of neuroinflammation in cognitive impairment is increasingly recognized. Non-steroidal anti-inflammatory drugs had been proven that it could improve cognitive impairment in large dose but with more side effect, which limited the application. The main objective of this study was to investigate whether the combined use of nicotine and celecoxib could obtain synergistic neuroprotective effect in ischemic rats. METHODS: Twenty adult Sprague-Dawley (SD) rats underwent ischemic model surgery by injecting endothelin-1 into the left thalamus, which were classified into four groups with different interventions: nicotine (1.5 mg/kg/d), celecoxib (15 mg/kg/d), nicotine (1.5 mg/kg/d) +celecoxib (15 mg/kg/d), or saline after surgery. The other five SD rats also underwent same surgery by injecting saline instead of endothelin-1, as the control group. Morris water maze (MWM) test was adopted to assess the cognition. Micro PET/CT with 2-[18F]-A-85380 were performed for α4ß2-nAChRs detection in vivo. Western blot, real-time PCR and immunohistochemical staining were adopted to detect the expression of α4ß2-nAChRs and inflammatory factors which included TNF-α, IL-1ß, IL-6 in brain tissue. Microglial activation in the brain was monitored by immunofluorescence with IBA1 staining. RESULTS: The MWM test showed rats given with nicotine or celecoxib alone showed much better memory than rats with saline, no difference was observed between nicotine and celecoxib. The rat memory was recovered most significant when the nicotine and celecoxib were combined (p < 0.05). Micro-PET/CT showed much more tracer uptake in the left thalamus and whole brain in rats given with nicotine, or nicotine + celecoxib (nico + cele group) than saline treated rats, whereas the rats given celecoxib did not. Compared with saline treated rats, we found the proteins of α4nAChR and ß2nAChR in rats given nicotine or nico + cele increased significantly, and mRNA/proteins of TNF-α, IL-1ß and IL-6 decreased at the same time. The α 4nAChR and ß 2nAChR proteins in rats given celecoxib is the same as saline treated rats, whereas the inflammatory factors decreased obviously compared with saline treated rats. Microglial activation was confirmed in saline treated rats, which was inhibited in rats give nicotine, celecoxib or both. CONCLUSIONS: The study revealed the combined use of nicotine and celecoxib may improve the cognitive function in ischemic rats, with a better effect than either alone. Both nicotine and celecoxib can inhibit inflammation, but through different mechanisms: nicotine can activate α4ß2-nAChRs while celecoxib is cyclooxygenase-2 inhibitor. Our findings suggest the combined application of two drugs with different anti-inflammation mechanism could attenuate cognitive impairment more effectively in ischemic rats, which may hold therapeutic potential in the clinical practice.

Anti-hyperalgesic effect of (-)-α-bisabolol and (-)-α-bisabolol/ß-Cyclodextrin complex in a chronic inflammatory pain model is associated with reduced reactive gliosis and cytokine modulation.

Chronic pain is a continuous or recurring pain which exceeds the normal course of recovery to an injury or disease. According to the origin of the chronic pain, it can be classified as inflammatory or neuropathic. This study aimed to evaluate the antinociceptive and anti-inflammatory effect of (-)-α-bisabolol (BIS) alone and complexed with ß-cyclodextrin (ßCD) in preclinical models of chronic pain. Chronic pain was induced by Freund's Complete Adjuvant (FCA) or partial lesion of the sciatic nerve (PLSN). Swiss mice were treated with BIS, BIS-ßCD (50 mg/kg, p.o) or vehicle (control) and mechanical hyperalgesia, thermal hyperalgesia, muscle strength and motor coordination were evaluated. In addition, levels of TNF-α and IL-10 and expression of the ionized calcium-binding adapter protein (IBA-1) were assessed in the spinal cord of the mice. The complexation efficiency of BIS in ßCD was evaluated by High-Performance Liquid Chromatography. BIS and BIS-ßCD reduced (p < 0.001) mechanical and thermal hyperalgesia. No alterations were found in force and motor coordination. In addition, BIS and BIS-ßCD inhibited (p < 0.05) TNF-α production in the spinal cord and stimulated (p < 0.05) the release of IL-10 in the spinal cord in PLSN-mice. Further, BIS and BIS-ßCD reduced IBA-1 immunostaining. Therefore, BIS and BIS-ßCD attenuated hyperalgesia, deregulated cytokine release and inhibited IBA-1 expression in the spinal cord in the PLSN model. Moreover, our results show that the complexation of BIS in ßCD reduced the therapeutic dose of BIS. We conclude that BIS is a promising molecule for the treatment of chronic pain.

Methamphetamine toxicity-induced calcineurin activation, nuclear translocation of nuclear factor of activated T-cells and elevation of cyclooxygenase 2 levels are averted by calpastatin overexpression in neuroblastoma SH-SY5Y cells.

Methamphetamine (METH) is an addictive stimulant drug that has many negative consequences, including toxic effects to the brain. Recently, the induction of inflammatory processes has been identified as a potential contributing factor to induce neuronal cell degeneration. It has been demonstrated that the expression of inflammatory agents, such as cyclooxygenase 2 (COX-2), depends on the activation of calcineurin (CaN) and nuclear factor of activated T-cells (NFAT). Moreover, the excessive elevation in cytosolic Ca2+ levels activates the cell death process, including calpain activation in neurons, which was diminished by the overexpression of the calpain inhibitor protein, calpastatin. However, it is unclear whether calpain mediates CaN-NFAT activation in the neurotoxic process. In the present study, we observed that the toxic high dose of METH-treated neuroblastoma SH-SY5Y cells significantly decreased cell viability but increased apoptotic cell death, the active cleaved form of calcineurin, the nuclear translocation of NFAT, and COX-2 levels. Nevertheless, these toxic effects were diminished in METH-treated calpastatin-overexpressing SH-SY5Y cells. These findings might emphasize the role of calpastatin against METH-induced toxicity by a mechanism related to calpain-dependent CaN-NFAT activation-induced COX-2 expression.

High Concentration of Calcium Promotes Mineralization in NRK-52E Cells Via Inhibiting the Expression of Matrix Gla Protein.

OBJECTIVE: To address whether matrix Gla protein (MGP) can inhibit mineralization in normal rat kidney tubular cells (NRK-52E) under high concentration of calcium. MATERIALS AND METHODS: NRK-52E cells were treated with high concentration of calcium. The viability and apoptosis of cells were detected by cell counting kit-8 and flow cytology, respectively. Real-time-polymerase chain, Western blotting, and immunofluorescence analysis were conducted to detect the expression of MGP. Cells were transfected with plasmid-MGP or siRNA-MGP for up- or down-regulation of the expression of MGP, respectively. Rat recombinant MGP was also used as supplementation of exogenous MGP. Alizarin red staining was conducted to detect the adherent and deposition of calcium salt. RESULTS: High concentration of calcium suppressed MGP expression in NRK-52E cells. There was significant mineralization when NRK-52E cells were treated with high concentration of calcium. Supplementation with exogenous rat recombinant MGP and overexpression of endogenous MGP both decreased the adherent and deposition of calcium salt to NRK-52E cells, while silence of MGP showed reverse results. CONCLUSION: MGP plays an inhibitory role in the stone formation. However, high concentration of calcium significantly inhibits the expression of MGP and then promotes mineralization in NRK-52E cells.

Endocrine sensitivity of estrogen receptor-positive breast cancer is negatively correlated with aspartate-ß-hydroxylase expression.

Although prognostic markers for early estrogen receptor (ER)-positive breast cancer have been extensively developed, predictive markers for adjuvant endocrine therapy are still lacking. Focusing on the mechanisms underlying endocrine resistance, we investigated whether the endocrine sensitivity of ER-positive breast cancer cells was correlated with the expression of aspartate-ß-hydroxylase (ASPH), which is involved in the development of hepatocellular carcinoma. ASPH expression in ER-positive and tamoxifen-resistant breast cancer cells was upregulated by the MAPK and phosphoinositide-3 kinase (PI3K) pathways, which both play pivotal roles in endocrine resistance. In the clinical setting, ASPH expression was negatively correlated with recurrence-free survival of luminal B breast cancer patients that received adjuvant endocrine therapy, but not in patients that did not receive adjuvant endocrine therapy. Luminal B breast cancer is one of the intrinsic molecular subtypes identified by the Prediction Analysis of Microarray 50 (PAM50) multiple gene classifier, and because of its poor response to endocrine therapy, chemotherapy in addition to endocrine therapy is generally required after surgical resection. Our results suggest that the endocrine sensitivity of luminal B breast cancer can be assessed by examining ASPH expression, which promotes the consideration of a prospective study on the association between ASPH expression at the mRNA and protein levels in luminal B breast cancer and subsequent response to endocrine therapy.

A survey of neuroimmune changes in pregnant and postpartum female rats.

During pregnancy and the postpartum period, the adult female brain is remarkably plastic exhibiting modifications of neurons, astrocytes and oligodendrocytes. However, little is known about how microglia, the brain's innate immune cells, are altered during this time. In the current studies, microglial density, number and morphological phenotype were analyzed within multiple regions of the maternal brain that are known to show neural plasticity during the peripartum period and/or regulate peripartum behavioral changes. Our results show a significant reduction in microglial density during late pregnancy and the early-mid postpartum period in the basolateral amygdala, medial prefrontal cortex, nucleus accumbens shell and dorsal hippocampus. In addition, microglia numbers were reduced postpartum in all four brain regions, and these reductions occurred primarily in microglia with a thin, ramified morphology. Across the various measures, microglia in the motor cortex were unaffected by reproductive status. The peripartum decrease in microglia may be a consequence of reduced proliferation as there were fewer numbers of proliferating microglia, and no changes in apoptotic microglia, in the postpartum hippocampus. Finally, hippocampal concentrations of the cytokines interleukin (IL)-6 and IL-10 were increased postpartum. Together, these data point to a shift in the maternal neuroimmune environment during the peripartum period that could contribute to neural and behavioral plasticity occurring during the transition to motherhood.

Treatment with tanshinone IIA suppresses disruption of the blood-brain barrier and reduces expression of adhesion molecules and chemokines in experimental autoimmune encephalomyelitis.

Tanshinone IIA (TSIIA), one of the major bioactive components of the traditional Chinese herb Salvia miltiorrhiza, has been reported to have both anti-inflammatory and immunoregulatory effects. The effect of treatment with TSIIA in multiple sclerosis, an autoimmune inflammatory neurodegenerative disease, however, remains poorly understood. In the present study, experimental autoimmune encephalomyelitis (EAE), a classical experimental model of MS, was used to investigate the therapeutic effect of TSIIA. TSIIA attenuated motor dysfunction and improved inflammation and demyelination associated with EAE in a dose-dependent manner. TSIIA also significantly reduced the levels of glial fibrillary acidic protein (GFAP) and ionized calcium-binding adapter molecule-1 (Iba-1), and protected the integrity of the blood-brain barrier (BBB) by increasing the expression of critical endothelial tight junction (TJ) proteins. TSIIA also inhibited the expression of some adhesion molecules and chemokines, which are considered to be critical for adhesion of immune cells and migration across the BBB. TSIIA was thus shown to be effective in the treatment of EAE through preventing the infiltration of immune cells into the CNS, strengthening the integrity of the BBB and decreasing the numbers of adhesion molecules and chemokines.

Biological therapy induces expression changes in Notch pathway in psoriasis.

Psoriasis is a chronic inflammatory skin disease, characterized by hyperproliferation of keratinocytes and by skin infiltration of activated T cells. To date, the pathophysiology of psoriasis has not yet been fully elucidated. The Notch pathway plays a determinant role in cell fate determination, proliferation, differentiation, immune cell development and function. Many biological agents, used in the treatment of psoriasis, include TFN-α inhibitors, such as etanercept, adalimumab, and anti IL-12/IL-23 p40 antibody, such as ustekinumab. This study aimed to determine mRNA expression levels by real-time RT-PCR, and protein expression levels, analysed by Western blot and immunohistochemistry, of some components of the Notch pathway, such as NOTCH1, NOTCH2, JAGGED1, and HES1 after biological treatments in psoriatic patients. mRNA and protein levels of NOTCH1, NOTCH2, JAGGED1 and HES1 were upregulated in skin samples from untreated psoriatic patients compared with normal controls. Biological therapy showed to downregulate differently the protein expression levels of the molecules under study. Our study suggests that Notch pathway components might be a potential therapeutic target against psoriasis.

Retinoic acid differentially affects in vitro proliferation, differentiation and mineralization of two fish bone-derived cell lines: different gene expression of nuclear receptors and ECM proteins.

Retinoic acid (RA), the main active metabolite of vitamin A, regulates vertebrate morphogenesis through signaling pathways not yet fully understood. Such process involves the specific activation of retinoic acid and retinoid X receptors (RARs and RXRs), which are nuclear receptors of the steroid/thyroid hormone receptor superfamily. Teleost fish are suitable models to study vertebrate development, such as skeletogenesis. Cell systems capable of in vitro mineralization have been developed for several fish species and may provide new insights into the specific cellular and molecular events related to vitamin A activity in bone, complementary to in vivo studies. This work aims at investigating the in vitro effects of RA (0.5 and 12.5 µM) on proliferation, differentiation and extracellular matrix (ECM) mineralization of two gilthead seabream bone-derived cell lines (VSa13 and VSa16), and at identifying molecular targets of its action through gene expression analysis. RA induced phenotypic changes and cellular proliferation was inhibited in both cell lines in a cell type-dependent manner (36-59% in VSa13 and 17-46% in VSa16 cells). While RA stimulated mineral deposition in VSa13 cell cultures (50-62% stimulation), it inhibited the mineralization of extracellular matrix in VSa16 cells (11-57% inhibition). Expression of hormone receptor genes (rars and rxrs), and extracellular matrix-related genes such as matrix and bone Gla proteins (mgp and bglap), osteopontin (spp1) and type I collagen (col1a1) were differentially regulated upon exposure to RA in proliferating, differentiating and mineralizing cultures of VSa13 and VSa16 cells. Altogether, our results show: (i) RA affects proliferative and mineralogenic activities in two fish skeletal cell types and (ii) that during phenotype transitions, specific RA nuclear receptors and bone-related genes are differentially expressed in a cell type-dependent manner.

S100P enhances the chemosensitivity of human gastric cancer cell lines.

BACKGROUND: The effect of the protein S100P on biological characteristics of cancer is not clear, especially in gastric cancer. We previously showed that S100P positive gastric cancer patients have a better cumulative survival than S100P negative patients. OBJECTIVE: To study the possible mechanisms of S100P enhanced the chemosensitivity to oxaliplatin in gastric cancer cell lines. METHODS: S100P was overexpressed in vitro by plasmid transfection and downregulated by siRNA transfection in the BGC823 and SGC7901 gastric cancer cell lines. Cell survival rate, changes in the chemoresistance gene, such as GST-π, MDR1, MRP1, Topo-II, MVP and BCRP, intake of anticancer drug were measured after oxaliplatin treatment. RESULTS: In SGC7901 cells, MTT assay indicated that increased S100P expression levels decreased the survival rate and decreased S100P expression levels increased the survival rate. In BGC823 and SGC7901 cell lines, mRNA of MDR1, a chemoresistance genes, was decreased in cells that overexpressed S100P, and increased in cells with downregulation of S100P. Intracellular accumulation of platinum increased in cells with overexpressed S100P, and decreased in cells with S100P downregulation. CONCLUSIONS: S100P contributes to oxaliplatin chemosensitivity in gastric cell lines by increasing drug inflow. It might also be a novel independent prognostic factor in gastric cancer patients who receive adjuvant chemotherapy with oxaliplatin.

Mammary analog secretory carcinoma of salivary glands: a report of 2 cases with expression of basal/myoepithelial markers (calponin, CD10 and p63 protein).

Mammary analog secretory carcinoma (MASC) of salivary glands is a recently described neoplasm with favorable outcome. We describe 2 cases of MASC occurring in a 34-year-old female and a 58-year-old male, both presenting with a swelling of upper lip and right parotid gland, measuring 15 and 20mm, respectively. Without adjuvant treatment, both patients have been free of disease for 15 months and 12 months since the operation. Microscopically, both tumors were cystic and showed tubular and cystopapillary architecture. The tumor cells had round to oval nuclei and eosinophilic cytoplasm. Presence of eosinophilic material was evident within cystic spaces. Immunohistochemically, both tumors expressed cytokeratins (CK), CK7, CK8, CK18, epithelial membrane antigen, vimentin, S-100 protein, mammaglobin, and STAT5a (signal transducer and activator of transcription 5a). Interestingly, both tumors showed variable expression of basal/myoepithelial markers. In one case, we observed diffuse expression of calponin and focal expression of p63 whereas expression of CD10 was absent. In the second case, the staining of calponin was negative, but there was focal expression of both p63 and CD10. Both neoplasms harbored the ETV6-NTRK3 fusion transcript as proved by RT-PCR. Although previously reported only rarely, we conclude that MASC may show expression of basal/myoepithelial markers.

Enhanced expression of nesfatin/nucleobindin-2 in white adipose tissue of ventromedial hypothalamus-lesioned rats.

Nesfatin-1, an anorexigenic protein, is ubiquitously expressed in the body. However, the exact mechanism underlying the in vivo regulation of production of nesfatin/nucleobindin-2 (NUCB2), a precursor protein of nesfatin-1, is unknown. We investigated the influence of modulation of autonomic nerve activity by a ventromedial hypothalamus (VMH) lesion and the subsequent effect on nesfatin/NUCB2 production in rat tissues innervated by the peripheral nervous system. Nesfatin/NUCB2 is strongly expressed in the pancreas and liver, moderately expressed in subcutaneous and visceral fat tissues and interscapular brown adipose tissue (iBAT), but is weakly expressed in the skeletal muscles. Our study results showed that the VMH lesion in VMH-lesioned rats did not affect nesfatin/NUCB2 expression in the pancreas, liver, skeletal muscle, and iBAT; however, the protein expression was significantly high in both subcutaneous and visceral fat tissues. In addition, continuous peripheral administration of carbachol for 5 days did not affect nesfatin/NUCB2 expression, but chemical sympathectomy using 6-hydroxydopamine mimicked the effect of VMH lesion by showing significantly high nesfatin/NUCB2 expression in the subcutaneous fat tissues. These results show that VMH lesion can modulate the autonomic nervous system activity and balance and increase nesfatin/NUCB2 expression in white adipose tissues of rats. Further, this action may be mediated via inhibition of the sympathetic nerve activity.

Insulin resistance induces medial artery calcification in fructose-fed rats.

Osteogenic differentiation of vascular smooth muscle cells (VSMCs) results in medial artery calcification, which is common in diabetes, but the pathogenesis is poorly understood. We aimed to explore the pathophysiological roles of insulin resistance (IR) on medial artery calcification in rats with 10% fructose in drinking water. After 12 weeks of fructose feeding, rats showed severe IR, with increased levels of fasting blood glucose, serum insulin and oral glucose tolerance test (OGTT). Fructose-fed rats showed aortic calcification, increased aortic calcium deposition and irregular elastic fibers in the medial layer of the vessel wall. Moreover, plasma phosphorus concentration, calcium × phosphorus product and alkaline phosphatase (ALP) activity, and aortic calcium content and ALP activity were significantly increased. Fructose feeding increased mRNA levels of osteopontin, type III sodium-dependent phosphate co-transporter, bone morphogenetic protein-2 and the key transcription factor core binding factor alpha 1 in aortic tissue and downregulated mRNA levels of osteoprotegerin and matrix γ-carboxyglutamic acid protein. Fructose feeding decreased protein levels of smooth-muscle lineage markers and induced severe lipid peroxidation injury. IR induced by high fructose feeding could evoke osteogenic transdifferentiation of VSMCs and promote vascular calcification.

Spatio-temporal expression analysis of the calcium-binding protein calumenin in the rodent brain.

Calumenin is a Ca(2+)-binding protein that belongs to the CREC superfamily. It contains six EF-hand domains that exhibit a low affinity for Ca(2+) as well as an endoplasmic reticulum retention signal. Calumenin exhibits a broad and relatively high expression in various brain regions during development as demonstrated by in situ hybridization. Signal intensity of calumenin is highest during the early development and then declines over time to reach a relatively low expression in adult animals. Immunohistochemistry indicates that at the P0 stage, calumenin expression is most abundant in migrating neurons in the zones around the lateral ventricle. In the brain of adult animals, it is expressed in various glial and neuronal cell types, including immature neurons in subgranular zone of hippocampal dentate gyrus. At the subcellular level, calumenin is identified in punctuate and diffuse distribution mostly in somatic regions where it co-localizes with endoplasmic reticulum (ER) and partially Golgi apparatus. Upon subcellular fractionation, calumenin is enriched in fractions containing membranes and is only weakly present in soluble fractions. This study points to a possible important role of calumenin in migration and differentiation of neurons, and/or in Ca(2+) signaling between glial cells and neurons.

17ß-Estradiol attenuates saturated fatty acid diet-induced liver injury in ovariectomized mice by up-regulating hepatic senescence marker protein-30.

Senescence marker protein-30 (SMP30) plays an important role in intracellular Ca(2+) homeostasis. The aim of the present study was to investigate the effects of estrogens on liver apoptotic damage and changes in SMP30 expression induced by a high saturated fatty acid diet (HSFD). Ovariectomized mice (OVX) and sham-operated mice (SHAM) were randomly divided into five groups: SHAM fed a normal diet (SHAM/ND), SHAM fed HSFD (SHAM/HSFD), OVX fed ND (OVX/ND), OVX fed HSFD (OVX/HSFD) and OVX fed HSFD with 17ß-estradiol (E2) supplementation using an implanted slow-release pellet (OVX/HSFD+E2). After 8 weeks, markers of endoplasmic reticulum (ER) stress and apoptosis, and levels of tumor necrosis factor-α (TNFα and SMP30 expression were investigated. Compared with SHAM/ND, OVX/HSFD mice showed significantly increased spliced X-box protein-1 (s-XBP1), phosphorylated eukaryotic initiation factor-2α (p-eIF2α), glucose-regulated protein 78 (GPR78), C/EBP homologous protein (CHOP), cytosolic cytochrome c, caspase-3 activity, and TNFα, and significantly decreased SMP30. These differences in OVX/HSFD mice were restored to the levels of SHAM/ND mice by E2 supplementation. These results suggest that E2 supplementation attenuates HSFD-induced liver apoptotic death in ovariectomized mice by up-regulating SMP30.

Spirulina platensis protects neurons via suppression of glial activation and peripheral sensitization leading to restoration of motor function in collagen-induced arthritic rats.

Spirulina platensis treatment (400 mg kg(-1) for 25 days) effectively suppressed peripheral sensitization via modulation of glial activation and improved motor coordination and restoration of functional motor activity in collagen-induced arthritic rats. Spirulina treatment also resulted in an appreciable reduction of the NF200 accumulation in the spinal cord neurons of arthritic rats. This is indicative of neuroprotective action of S. platensis against glutamate excitotoxicity-induced central sensitization produced by the peripheral joint inflammation in the collagen-induced arthritis. The results suggest that effects of S. platensis may be due to its counter regulation of spinal glial activation and could be a potential strategy for the treatment of arthritis.

Myocardial transfection with naked DNA plasmid encoding hepatocyte growth factor prevents the progression of heart failure in dogs.

This study examined the effects of localized intramyocardial injections of hepatocyte growth factor (HGF) naked DNA plasmid on the progression of left ventricular (LV) dysfunction and remodeling in dogs with moderate heart failure (HF). Twenty-one dogs with intracoronary microembolization-induced HF [LV ejection fraction (EF) = 35-40%] were randomized into three treatment groups, namely, high-dose HGF plasmid (4.0 mg, n = 7), low-dose HGF plasmid (0.4 mg, n = 7), and sham-operated controls treated with normal saline (n = 7). A total of 10-15 injections of HGF plasmid or saline were made directly into the anterior wall of LV. LV EF and end-systolic volume (ESV) were measured before randomization (pretreatment) and at the end of 3 mo of follow-up (posttreatment). Treatment effect (Δ) was calculated as the change from pre- to posttreatment. Protein expression of sarcoplasmic reticulum (SR) Ca(2+)-cycling proteins was determined in LV tissue obtained from the sites of HGF injection and remote areas. Low-dose HGF attenuated the decline in EF (ΔEF: -3 ± 1 vs. -8 ± 1%, P < 0.05) and the increase in ESV (ΔESV: 6 ± 2 vs. 10 ± 1 ml, P < 0.05) seen in control sham-operated dogs, whereas high-dose HGF significantly increased EF (ΔEF: 4 ± 1 vs. -8 ± 1%, P < 0.05) and prevented the increase in ΔESV (ESV: -1 ± 1 vs. 10 ± 1 ml, P < 0.05) compared with control dogs. Treatment with high- and low-dose HGF improved the expression of the SR Ca(2+)-cycling proteins compared with controls. In conclusion, regional intramyocardial injections of HGF naked DNA plasmid improve regional and global LV function and prevent progressive LV remodeling.

Molecular characterization of the song control nucleus HVC in Bengalese finch brain.

Songbirds have a specialized neural substrate for learned vocalization, called the song circuit, which consists of several song nuclei in the brain. The song control nucleus HVC (a letter-based name) is the intersection point of the song learning and vocal motor pathways. Knowledge of the types of genes expressed in the HVC is essential in understanding the molecular aspects of the HVC. Gene expression in the HVC under silent conditions shows the competence necessary for singing. To investigate this, we compared the HVC with its adjacent tissues in searching for the molecular specificities of the song nucleus HVC using an in-house cDNA microarray of the Bengalese finch (Lonchura striata var. domestica). Our microarray analysis revealed that 70 genes were differentially expressed in the HVC compared with the adjacent tissue. We investigated 27 of the microarray-selected genes that were enriched or repressed in the HVC by in situ hybridization. We found that multiple calcium-binding proteins (e.g., CAPS2, parvalbumin and ATH) were enriched in the HVC. Meanwhile, the adult HVC showed low expression levels of plasticity-related genes (e.g., CAMK2A and MAP2K1) compared with the juvenile HVC. The HVC plays an important role during song learning, but our results suggest that the plasticity of this nucleus may be suppressed during adulthood. Our findings provide new information about the molecular features that characterize the HVC.

Characterisation and potential diagnostic value of circulating matrix Gla protein (MGP) species.

Matrix γ-carboxyglutamate (Gla) protein (MGP) is an important local inhibitor of vascular calcification, which can undergo two post-translational modifications: vitamin K-dependent γ-glutamate carboxylation and serine phosphorylation. While carboxylation is thought to have effects upon binding of calcium-ions, phosphorylation is supposed to affect the cellular release of MGP. Since both modifications can be exerted incompletely, various MGP species can be detected in the circulation. MGP levels were measured with two commercially available competitive and two novel sandwich assays in healthy controls, in patients with rheumatic disease, aortic valve disease, and end-stage renal disease, as well as in volunteers after vitamin K supplementation (VKS) and treatment with vitamin K antagonists (VKA). Major differences were found between the MGP assays, including significantly different behaviour with regard to vascular disease and the response to VKA and VKS. The dual-antibody assay measuring non-phosphorylated, non-carboxylated MGP (dp-ucMGP) was particularly sensitive for these changes and would be suited to assess the vascular vitamin K status. We conclude that the different assays for particular circulating MGP species allows the assessment of various aspects of the MGP system.