Vitamin K antagonist anticoagulant usage is associated with increased incidence and progression of osteoarthritis.

OBJECTIVES: Vitamin K is hypothesised to play a role in osteoarthritis (OA) pathogenesis through effects on vitamin K-dependent bone and cartilage proteins, and therefore may represent a modifiable risk factor. A genetic variant in a vitamin K-dependent protein that is an essential inhibitor for cartilage calcification, matrix Gla protein (MGP), was associated with an increased risk for OA. Vitamin K antagonist anticoagulants (VKAs), such as warfarin and acenocoumarol, act as anticoagulants through inhibition of vitamin K-dependent blood coagulation proteins. VKAs likely also affect the functioning of other vitamin K-dependent proteins such as MGP. METHODS: We investigated the effect of acenocoumarol usage on progression and incidence of radiographic OA in 3494 participants of the Rotterdam Study cohort. We also examined the effect of MGP and VKORC1 single nucleotide variants on this association. RESULTS: Acenocoumarol usage was associated with an increased risk of OA incidence and progression (OR=2.50, 95% CI=1.94-3.20), both for knee (OR=2.34, 95% CI=1.67-3.22) and hip OA (OR=2.74, 95% CI=1.82-4.11). Among acenocoumarol users, carriers of the high VKORC1(BB) expression haplotype together with the MGP OA risk allele (rs1800801-T) had an increased risk of OA incidence and progression (OR=4.18, 95% CI=2.69-6.50), while this relationship was not present in non-users of that group (OR=1.01, 95% CI=0.78-1.33). CONCLUSIONS: These findings support the importance of vitamin K and vitamin K-dependent proteins, as MGP, in the pathogenesis of OA. Additionally, these results may have direct implications for the clinical prevention of OA, supporting the consideration of direct oral anticoagulants in favour of VKAs.

Acrylamide treatment alters the level of Ca2+ and Ca2+-related protein kinase in spinal cords of rats.

This study aimed to analyze the neurological changes induced by acrylamide (ACR) poisoning and their underlying mechanisms within the spinal cords of male adult Wistar rats. The rats were randomly divided into three groups (n = 9 rats per group). ACR was intraperitoneally injected to produce axonopathy according to the daily dosing schedules of 20 or 40 mg/kg/day of ACR for eight continuous weeks (three times per week). During the exposure period, body weights and gait scores were assessed, and the concentration of Ca2+ was calculated in 27 mice. Protein kinase A (PKA), protein kinase C (PKC), cyclin-dependent protein kinase 5 (CDK5), and P35 were assessed by electrophoretic resolution and Western blotting. The contents of 3'-cyclic adenosine monophosphate (cAMP) and calmodulin (CaM) were determined using ELISA kits, and the activities of calcium/calmodulin-dependent protein kinase II (CaMKII), PKA, and PKC were determined using the commercial Signa TECTPKAassay kits. Compared with control rats, treatment with 20 and 40 mg/kg of ACR decreased body weight and increased gait scores at 8 weeks. Intracellular Ca2+ levels increased significantly in treated rats; CaM, PKC, CDK5, and P35 levels were significantly decreased; and PKA and cAMP levels remained unchanged. CaMKII, PKA, and PKC activities increased significantly. The results indicated that ACR can damage neurofilaments by affecting the contents and activities of CaM, CaMKII, PKA, cAMP, PKC, CDK5, and P35, which could result in ACR toxic neuropathy.

Effect of Berberine Isolated from Barberry Species by Centrifugal Partition Chromatography on Memory and the Expression of Parvalbumin in the Mouse Hippocampus Proper.

Neurodegenerative diseases associated with memory disturbances are important health issues occurring due to a prolonged life span. This article presents the results of a study targeting the emergence of a drug candidate with antiamnesic properties. The effect of berberine (BBR), an isoquinoline alkaloid isolated from the overground parts of Berberis sibirica Pall., on memory and expression of parvalbumin in the mouse hippocampus proper were determined. High-purity BBR was isolated by centrifugal partition chromatography from a methanolic extract from B. sibirica by using a methyl-tert-butyl ether and water (1:1 v/v) solvent system with 10 mmol/L of triethylamine and hydrochloric acid. In an in vivo study, we assessed the influence of the chronic administration of BBR on different stages of memory-related responses in mice. Our results indicated that the chronic administration of BBR in a higher dose (5 mg/kg) improves long-term memory acquisition in mice, as determined in the passive avoidance test. The hippocampal CA1-CA3 fields showed an increased number of parvalbumin-immunoreactive neurons (PV-IR) and nerve fibers as compared to the control. No significant changes in the dentate gyrus were observed between the groups. The HPLC-ESI-QTOF-MS/MS analysis of the biological material revealed the content of BBR as 363.4 ± 15.0 ng (4.11% of RSD) per brain, 15.06 ± 0.89 ng (5.91% of RSD) per hippocampus, and 54.45 ± 1.40 (4.05% of RSD) ng in 100 µL plasma. The study showed that BBR could be a factor influencing the expression of PV in hippocampal neurons. We speculate that BBR may modulate the level of Ca2+ in neurons and thus potentially act as a neuroprotective factor against neuronal damages.

Vitamin K1 Inhibition of Renal Crystal Formation through Matrix Gla Protein in the Kidney.

BACKGROUND AND OBJECTIVES: Vitamin K (VK) plays a major role in modifying the binding of calcium in bones and blood vessels. Understanding the effect of VK on crystal formation in the kidney would contribute to advancing the treatment and prevention of kidney stones. METHODS: Rats were treated with vitamin K1 (VK1) for 8 weeks. VK1 levels were detected and crystal formation were observed. HK2 cells were exposed to calcium oxalate monohydrate crystals. Apoptosis and cell viability were detected. Crystal deposition was analyzed using atomic absorption assay. The adenovirus vectors expressing matrix Gla protein (MGP) and siMGP were constructed to elucidate the effect and mechanism of VK1 on crystal formation. MGP expression in vivo and in vitro was analyzed by Western blot. The mRNA levels of monocyte chemoattractant protein-1 (MCP-1) and collagen I was measured by semiquantitative RT-PCR. RESULTS: The concentrations of VK1 in whole blood and kidney tissues rose under treatment with VK1. Crystal formation was inhibited from the second to the 6th week, the frequency and quality of crystal formation decreased significantly, and the location of crystal formation was limited to a greater extent in the rats treated by VK1 compared to the control group. Warfarin treatment in the crystals-exposed HK2 cells significantly increased the number of crystals adhering to cells and the number of apoptotic cells and reduced cell viability. VK1 treatment reversed warfarin's above influence. VK1 inhibited the upregulations of MCP-1 and collagen I in kidney tissues under crystal load. VK1 treatment increased MGP expression in vivo and in vitro, and MGP is necessary for VK1 to play a role in crystal deposition in cells. CONCLUSIONS: VK1 treatment can inhibit the formation of renal crystals in vivo. VK1 increases MGP expression and functions through MGP to reduce crystal deposition in cells and provide cell protection. Our findings suggest that VK1 treatment could be a potential strategy for the treatment and prevention of nephrolithiasis.

The Differential Response to Ca2+ from Vertebrate and Invertebrate Calumenin Is Governed by a Single Amino Acid Residue.

Calumenin (Calu) is a well-conserved multi-EF-hand-containing Ca2+-binding protein. In this work, we focused on the alterations that calumenin has undergone during evolution. We demonstrate that vertebrate calumenin is significantly different from its invertebrate homologues with respect to its response to Ca2+ binding. Human calumenin (HsCalu1) is intrinsically unstructured in the Ca2+ free form and responds to Ca2+ with a dramatic gain in structure. Calumenin from Caenorhabditis elegans (CeCalu) is structured even in the apo form, with no conformational change upon binding of Ca2+. We decode this structural and functional distinction by identifying a single "Leu" residue-based switch located in the fourth EF-hand of HsCalu1, occupied by "Gly" in the invertebrate homologues. We demonstrate that replacing Leu with Gly (L150G) in HsCalu1 enables the protein to adopt a structural fold even in the Ca2+ free form, similar to CeCalu, leading to ligand compensation (adoption of structure in the absence of Ca2+). The fourth (of seven) EF-hand of HsCalu1 nucleates the structural fold of the protein depending on the switch residue (Gly or Leu). Our analyses reveal that the Leu that replaced Gly from fishes onward is absolutely conserved in higher vertebrates, while lower organisms have Gly, not only enlarging the scope of Ca2+-dependent structural transitions but also drawing a boundary between the invertebrate and vertebrate calumenin. The evolutionary selection of the switch residue strongly corroborates the change in the structure of the protein and its pleiotropic functions and seems like it can be extended to the presence or absence of a heart in that organism.

Neuroprotective effects of SMTP-44D in mice stroke model in relation to neurovascular unit and trophic coupling.

Stachybotrys microspora triprenyl phenol (SMTP)-44D has both anti-oxidative and anti-inflammatory activities, but its efficacy has not been proved in relation to the pathological changes of neurovascular unit (NVU) and neurovascular trophic coupling (NVTC) in ischemic stroke. Here, the present study was designed to assess the efficacies of SMTP-44D, moreover, compared with the standard neuroprotective reagent edaravone in ischemic brains. ICR mice were subjected to transient middle cerebral artery occlusion (tMCAO) for 60 min, SMTP-44D (10 mg/kg) or edaravone (3 mg/kg) was intravenously administrated through subclavian vein just after the reperfusion, and these mice were examined at 1, 3, and 7 d after reperfusion. Compared with the vehicle group, SMTP-44D treatment revealed obvious ameliorations in clinical scores and infarct volume, meanwhile, markedly suppressed the accumulations of 4-HNE, 8-OHdG, nitrotyrosine, RAGE, TNF-α, Iba-1, and cleaved caspase-3 after tMCAO. In addition, SMTP-44D significantly prevented the dissociation of NVU and improved the intensity of NAGO/BDNF and the number of BDNF/TrkB and BDNF/NeuN double positive cells. These effects of SMTP-44D in reducing oxidative and inflammatory stresses were similar to or stronger than those of edaravone. The present study demonstrated that SMTP-44D showed strong anti-oxidative, anti-inflammatory, and anti-apoptotic effects, moreover, the drug also significantly improved the NVU damage and NVTC in the ischemic brain.

Effects of Post-Treatment Hydrogen Gas Inhalation on Uveitis Induced by Endotoxin in Rats.

BACKGROUND Molecular hydrogen (H2) has been widely reported to have benefiicial effects in diverse animal models and human disease through reduction of oxidative stress and inflammation. The aim of this study was to investigate whether hydrogen gas could ameliorate endotoxin-induced uveitis (EIU) in rats. MATERIAL AND METHODS Male Sprague-Dawley rats were divided into a normal group, a model group, a nitrogen-oxygen (N-O) group, and a hydrogen-oxygen (H-O) group. EIU was induced in rats of the latter 3 groups by injection of lipopolysaccharide (LPS). After that, rats in the N-O group inhaled a gas mixture of 67% N2 and 33% O2, while those in the H-O group inhaled a gas mixture of 67% H2 and 33% O2. All rats were graded according to the signs of uveitis after electroretinography (ERG) examination. Protein concentration in the aqueous humor (AqH) was measured. Furthermore, hematoxylin-eosin staining and immunostaining of anti-ionized calcium-binding adapter molecule 1 (Iba1) in the iris and ciliary body (ICB) were carried out. RESULTS No statistically significant differences existed in the graded score of uveitis and the b-wave peak time in the Dark-adapted 3.0 ERG among the model, N-O, and H-O groups (P>0.05), while rats of the H-O group showed a lower concentration of AqH protein than that of the model or N-O group (P<0.05). The number of the infiltrating cells in the ICB of rats from the H-O group was not significantly different from that of the model or N-O group (P>0.05), while the activation of microglia cells in the H-O group was somewhat reduced (P<0.05). CONCLUSIONS Post-treatment hydrogen gas inhalation did not ameliorate the clinical signs, or reduce the infiltrating cells of EIU. However, it inhibited the elevation of protein in the AqH and reduced the microglia activation.

The effects of Desflurane and Sevoflurane on Nesfatin-1 levels in laparoscopic Cholecystectomy: a randomized controlled trial.

BACKGROUND: Nesfatin-1 is involved in cardiovascular regulation, stress-related responses. The objective of this study is to investigate the impact of volatile anesthetics on Nesfatin-1 levels. METHOD: Fourty-two patients aged 30-65 years with the American Society Anesthesiology (ASA) Class I-II who were scheduled for laparoscopic cholecystectomy were included in the study Patients were randomized into two group; desflurane administered group (Group I, n = 21) and sevoflurane administered group (Group II, n = 21). For anesthesia maintenance, the patients received 6% desflurane or 2% sevoflurane in 40% O2 and 60% air. The patient's heart rate (HR), mean, systolic and diastolic arterial pressures (MAP, SAP, DAP), peripheral O2 saturation (SpO2) were monitored and recorded before induction, after induction, after intubation, and during extubation. Blood samples were collected before induction (T1), and after extubation when aldrete score was 10 (T2). RESULTS: Demographic data were similar between the groups. The preoperative levels of nesfatin were similar in the two groups (p = 0.715). In desflurane group, post-operative nesfatin levels were similar compared to preoperative levels (p = 0.073). In sevoflurane group, post-operative nesfatin levels were similar (p = 0.131). The nesfatin levels (postoperative vs preoperative) were similar between the groups (p = 0.900). CONCLUSION: In conclusion, this study results suggest that nesfatin-1 levels are not affected by the use of sevoflurane or desflurane in patients undergoing laparoscopic cholecystectomy. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry, ACTRN12617001023347 , retrospectively registered on 17 July 2017.

Infliximab therapy reverses the increase of allograft inflammatory factor-1 in serum and colonic mucosa of rats with inflammatory bowel disease.

OBJECTIVE: Our purpose was to study the molecular basis of infliximab (IFX) effect on colon mucosa in a colitis model and to identify new biomarkers of mucosal healing. METHODS: Healthy rats and rats which were subjected to experimental colitis induced by dextran sulfate sodium, with or without IFX treatment (in the short- and long-term), were studied along with forty-seven IBD patients. Colon mucosal integrity by periodic acid Schiff (PAS) staining, intestinal damage by immunohistochemistry (proliferating cell nuclear antigen, ß-catenin, E-cadherin, phosphotyrosine, p-p38, allograft inflammatory factor-1 (AIF-1) and colonic mucosal apoptosis by TUNEL staining were evaluated in rats while serum and colon AIF-1 levels were determined in IBD patients. RESULTS: In rats with colitis, IFX reestablished the epithelial barrier integrity, recovered mucus production and decreased colon inflammation, as verified by reduced serum and colon AIF-1 levels; colon and serum AIF-1 levels were also lower in inactive IBD patients compare to active ones. P38 activation after IFX treatment tended to induce differentiation/proliferation of epithelial cells along the colonic crypt-villous axis. CONCLUSIONS: These findings support AIF-1 as a new biomarker of mucosal healing in experimental colitis and suggest that p38 activation is involved in the mucosal healing intracellular mechanism induced by IFX treatment.

Resveratrol protects against lipopolysaccharide-induced cardiac dysfunction by enhancing SERCA2a activity through promoting the phospholamban oligomerization.

The bacterial endotoxin lipopolysaccharide (LPS) is a main culprit responsible for cardiac dysfunction in sepsis. This study examined whether resveratrol could protect against LPS-induced cardiac dysfunction by improving the sarcoplasmic endoplasmic reticulum Ca2+-ATPase (SERCA2a) activity. Echocardiographic parameters, cardiomyocyte contractile and Ca2+ transient properties, markers for cardiac inflammation, cell death, and oxidative stress, SERCA2a activity, and the ratios of phospholamban (PLB) monomer to oligomer were measured. Cardiac function was decreased >50% after LPS challenge (6 mg/kg for 6 h), which was improved by resveratrol. There was neither difference in plasma tumor necrosis factor-α and troponin I levels nor in infiltration of CD45+ cells in cardiac tissue between resveratrol-treated and untreated groups. In cardiomyocytes, LPS significantly decreased contractile amplitude, elongated relengthening time, diminished Ca2+ transient, reduced SERCA2a activity, and increased superoxide generation. These pathological alterations were attenuated by resveratrol treatment. Immunoblot analysis showed that LPS-treated mice had increased levels of malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), and the monomer form of PLB, along with decreases in the levels of SERCA2a, the oligomer form of PLB and nuclear factor erythroid 2-related factor (Nrf-2). Resveratrol treatment upregulated SERCA2a, the oligomer form of PLB, and Nrf-2 expression and function, and downregulated MDA, 4-HNE, and the monomer form of PLB. Our data suggest that the activity of SERCA2a in endotoxemia is inhibited, possibly due to increases in the monomer form of PLB. Resveratrol protects the heart from LPS-induced injuries at least in part through promoting the oligomerization of PLB that leads to enhanced SERCA2a activity.

Plasma Desphospho-Uncarboxylated Matrix Gla Protein as a Marker of Kidney Damage and Cardiovascular Risk in Advanced Stage of Chronic Kidney Disease.

BACKGROUND/AIMS: Desphospho-uncarboxylated matrix Gla protein (dp-ucMGP) is formed as a result of vitamin K insufficiency. The aim of this study was to investigate the association between plasma dp-ucMGP, kidney function and cardiovascular risk factors before and after 9-months substitution of vitamin K2 in non-dialysis patients with chronic kidney disease (CKD) stage 4 and 5. METHODS: 38 CKD patients were supplemented for 270±12 days with 90 µg vitamin K2 and 10 µg cholecalciferol or 10 µg cholecalciferol alone. At baseline and at follow-up circulating calcium, phosphate, lipids, hemoglobin, albumin and total protein, dp-ucMGP, osteoprotegerin, fetuin A, osteocalcin and fibroblast grown factor 23 (FGF-23) were assessed. Proteinuria was assessed in the first morning void. RESULTS: Baseline plasma dp-ucMGP was 1018.6±498.3 pmol/l and was significantly higher in patients at stage 5 CKD (1388.3 ±505.4 pmol/l) than at stage 4 (885.1±419.7 pmol/l), p=0.04. Vitamin K2 supplementation resulted in a decrease of dp-ucMGP level by 10.7%. Plasma dp-ucMGP was positively associated with proteinuria, serum creatinine, PTH and FGF-23; and inversely associated with glomerular filtration rate, serum hemoglobin and albumin. CONCLUSIONS: High dp-ucMGP level, reflecting a poor vitamin K status seems to be associated with kidney damage and may be also a marker of cardiovascular risk in CKD patients. Supplementation with vitamin K2 may improve the carboxylation status of MGP.

Ultrasonography reveals in vivo dose-dependent inhibition of end systolic and diastolic volumes, heart rate and cardiac output by nesfatin-1 in zebrafish.

Nesfatin-1 is an 82 amino acid peptide that inhibits food intake in rodents and fish. While endogenous nesfatin-1, and its role in the regulation of food intake and hormone secretion has been reported in fish, information on cardiovascular functions of nesfatin-1 in fish is in its infancy. We hypothesized that cardiac NUCB2 expression is meal responsive and nesfatin-1 is a cardioregulatory peptide in zebrafish. NUCB2/nesfatin-1 like immunoreactivity was detected in zebrafish cardiomyocytes. Real-time quantitative PCR analysis found that the cardiac expression of NUCB2A mRNA in unfed fish decreased at 1h post-regular feeding time. Food deprivation for 7days did not change NUCB2A mRNA expression. However, NUCB2B mRNA expression was increased in the heart of zebrafish after a 7-day food deprivation. Ultrasonography of zebrafish heart at 15min post-intraperitoneal injection of nesfatin-1 (250 and 500ng/g body weight) showed a dose-dependent inhibition of end diastolic and end systolic volumes. A dose dependent decrease in heart rate and cardiac output was observed in zebrafish that received nesfatin-1, but no changes in stroke volume were found. Nesfatin-1 treatment caused a significant increase in the expression of Atp2a2a mRNA encoding the calcium-handling pump, SERCA2a, while it had no effects on the expression of calcium handling protein RyR1b encoding mRNA. Our data support cardiosuppressive effects of nesfatin-1 in zebrafish, and reveals energy availability as one determinant of cardiac NUCB2 mRNA expression.

N-acetylcysteine reverses diastolic dysfunction and hypertrophy in familial hypertrophic cardiomyopathy.

S-glutathionylation of cardiac myosin-binding protein C (cMyBP-C) induces Ca(2+) sensitization and a slowing of cross-bridge kinetics as a result of increased oxidative signaling. Although there is evidence for a role of oxidative stress in disorders associated with hypertrophic cardiomyopathy (HCM), this mechanism is not well understood. We investigated whether oxidative myofilament modifications may be in part responsible for diastolic dysfunction in HCM. We administered N-acetylcysteine (NAC) for 30 days to 1-mo-old wild-type mice and to transgenic mice expressing a mutant tropomyosin (Tm-E180G) and nontransgenic littermates. Tm-E180G hearts demonstrate a phenotype similar to human HCM. After NAC administration, the morphology and diastolic function of Tm-E180G mice was not significantly different from controls, indicating that NAC had reversed baseline diastolic dysfunction and hypertrophy in our model. NAC administration also increased sarco(endo)plasmic reticulum Ca(2+) ATPase protein expression, reduced extracellular signal-related kinase 1/2 phosphorylation, and normalized phosphorylation of phospholamban, as assessed by Western blot. Detergent-extracted fiber bundles from NAC-administered Tm-E180G mice showed nearly nontransgenic (NTG) myofilament Ca(2+) sensitivity. Additionally, we found that NAC increased tension cost and rate of cross-bridge reattachment. Tm-E180G myofilaments were found to have a significant increase in S-glutathionylation of cMyBP-C, which was returned to NTG levels upon NAC administration. Taken together, our results indicate that oxidative myofilament modifications are an important mediator in diastolic function, and by relieving this modification we were able to reverse established diastolic dysfunction and hypertrophy in HCM.

Analysis of the Leishmania peroxin 7 interactions with peroxin 5, peroxin 14 and PTS2 ligands.

LPEX7 (Leishmania peroxin 7) is essential for targeting newly synthesized proteins with a PTS2 (peroxisome-targeting signal type 2) import signal into the glycosome. In the present paper, we describe the biophysical characterization of a functional LPEX7 isolated from Escherichia coli inclusion bodies. Pull-down assays showed that LPEX7 binds the interacting partners LdPEX5 (Leishmania donovani peroxin 5) and LdPEX14, but, more importantly, this receptor can specifically bind PTS2 cargo proteins in the monomeric and dimeric states. However, in the absence of interacting partners, LPEX7 preferentially adopts a tetrameric structure. Mapping studies localized the LdPEX5- and LdPEX14-binding sites to the N-terminal portion of LPEX7. Deletion of the first 52 residues abolished LdPEX14 association without altering the LdPEX5 interaction. Intrinsic fluorescence techniques suggested that each LPEX7 subunit has a single unique binding site for each of the respective interacting partners LdPEX5, LdPEX14 and PTS2 cargo proteins. Extrinsic fluorescence studies with ANS (8-anilinonaphthalene-1-sulfonic acid) demonstrated that LPEX7 contains a surface-exposed hydrophobic region(s) that was not altered by the binding of a PTS2 protein or LdPEX5. However, in the presence of these ligands, the accessibility of the hydrophobic domain was dramatically restricted, suggesting that both ligands are necessary to induce notable conformational changes in LPEX7. In contrast, binding of LdPEX14 did not alter the hydrophobic domain on LPEX7. It is possible that the hydrophobic surfaces on LPEX7 may be a crucial characteristic for the shuttling of this receptor in and out of the glycosome.

Supplementation of apelin increase plasma levels of nesfatin-1 in normal and DOCA-salt hypertensive rats.

AIMS: We aimed to observe the effects of apelin supplementation on the plasma levels of nesfatin-1 in DOCA-salt hypertensive and normal rats. METHODS: For this purpose, 28 young Wistar albino male rats were divided into four groups; Control (C), Control + Apelin (C+A), Hypertension (HT) and Hypertension + Apelin (HT+A). Hypertension was induced by injection of DOCA-salt (25 mg/kg, s.c.) twice weekly, 4 weeks, whereas intraperitoneal apelin was administered (200 µg.kg-1) for 17 days. Plasma nesfatin-1 and apelin levels were measured with ELISA. Systolic blood pressure was monitored using a tail cuff system. The relationships between plasma nesfatin levels and blood pressure were assessed. RESULTS: Plasma nesfatin-1 levels was found lower in control animals compared to C+A, HT and HT+A groups (p = 0.002, p = 0.026 and p = 0.011, respectively). Systolic blood pressures were similar in the C and C+A groups, but systolic blood pressures of the HT and HT+A groups was found significantly higher than the C and C+A groups. CONCLUSIONS: In conclusion, apelin administration induced an increment of nesfatin-1 in normal rats and plasma levels of nesfatin-1 increase in DOCA-salt hypertension rats. But apelin addition in hypertension did not cause an extra increase in nesfatin-1 levels. This is the first report to investigate the effect of apelin administration on plasma nesfatin levels of normal and hypertensive rats (Fig. 2, Ref. 44).

FGF-23 dysregulates calcium homeostasis and electrophysiological properties in HL-1 atrial cells.

BACKGROUND: Fibroblast growth factor (FGF)-23 is a key regulator of phosphate homeostasis. Higher FGF-23 levels are correlated with poor outcomes in cardiovascular diseases. FGF-23 can produce cardiac hypertrophy and increase intracellular calcium, which can change cardiac electrical activity. However, it is not clear whether FGF-23 possesses arrhythmogenic potential through calcium dysregulation. Therefore, the purposes of this study were to evaluate the electrophysiological effects of FGF-23 and identify the underlying mechanisms. METHODS: Patch clamp, confocal microscope with Fluo-4 fluorescence, and Western blot analyses were used to evaluate the electrophysiological characteristics, calcium homeostasis and calcium regulatory proteins in HL-1 atrial myocytes with and without FGF-23 (10 and 25 ng/mL) incubation for 24 h. RESULTS: FGF-23 (25 ng/mL) increased L-type calcium currents, calcium transient and sarcoplasmic reticulum Ca(2+) contents in HL-1 cells. FGF-23 (25 ng/mL)-treated cells (n = 14) had greater incidences (57%, 17% and 15%, P < 0·05) of delayed afterdepolarizations than control (n = 12) and FGF-23 (10 ng/mL)-treated cells (n = 13). Compared with control cells, FGF-23 (25 ng/mL)-treated cells (n = 14) exhibited increased phosphorylation of calcium/calmodulin-dependent protein kinase IIδ and phospholamban (PLB) at threonine 17 but had similar phosphorylation extents of PLB at serine 16, total PLB and sarcoplasmic reticulum Ca(2+) -ATPase protein. Moreover, the FGF receptor inhibitor (PD173074, 10 nM), calmodulin inhibitor (W7, 5 µM) and phospholipase C inhibitor (U73122, 1 µM) attenuated the effects of FGF-23 on calcium/calmodulin-dependent protein kinase II phosphorylation. CONCLUSIONS: FGF-23 increases HL-1 cells arrhythmogenesis with calcium dysregulation through modulating calcium-handling proteins.

Effects of thymoquinone on STZ-induced diabetic nephropathy: an immunohistochemical study.

BACKGROUND: Thymoquinone (TQ) is the most abundant and active ingredient of Nigella sativa (NS) seeds. Its hepatic, renal, and cardiac protective effects have been demonstrated in animal models. Streptozotocin (STZ) is an antibiotic that is widely used experimentally as an agent capable of inducing insulin-dependent diabetes mellitus (IDDM), also known as type I diabetes mellitus (T1DM). OBJECTIVES: This study was carried out in an attempt to highlight the possible beneficial effects of TQ in STZ-induced diabetes in rats and to determine the predictive value of mesenchymal and epithelial markers in the response of diabetic nephropathy to TQ. MATERIALS AND METHODS: Sixty adult male albino rats were divided in 3 groups: control, diabetic untreated, and diabetic treated with TQ. RESULTS: Diabetic rats exhibited morphological changes in both renal glomeruli and tubules with immunohistochemical expression of the mesenchymal markers Fsp1, desmin, and MMP-17 and disappearance of the epithelial marker ZO-1 largely in the glomeruli of diabetic kidneys. Treatment with TQ significantly attenuated renal morphological and immunohistochemical changes in STZ-induced diabetic rats. CONCLUSIONS: Thymoquinone has protective effects on experimental diabetic nephropathy. Both mesenchymal and epithelial markers serve as excellent predictors of early kidney damage and indicators of TQ responsiveness in STZ-induced diabetic nephropathy.

GSI-I has a better effect in inhibiting hepatocellular carcinoma cell growth than GSI-IX, GSI-X, or GSI-XXI.

Current studies are ongoing to find new drugs for the treatment of hepatocellular carcinoma (HCC). The discovery of drugs depends on the identification of molecules that can play essential roles in the development of liver cancer, for example, Notch pathway molecules. γ-Secretase inhibitors (GSIs) can inhibit the cleavage of intramembranous substrates of all Notch receptors and subsequently suppress Notch signaling. However, whether the inhibition of the Notch pathway can suppress or promote HCC growth is still under debate. In this study, we examined the expression of Notch pathway molecules in 20 pairs of HCC tissue with their normal counterparts and a panel of eight HCC cell lines. We also determined the effects of different types of GSI treatments on the cell growth of those HCC cell lines. Our results showed that the molecules of the Notch pathway were expressed in six of the eight HCC cell lines. Those six HCC cell lines were more sensitive to GSI-I treatment than the nonexpression ones. Among the four inhibitors, GSI-X and GSI-XXI exerted no effect on HCC cells growth at all. GSI-IX inhibited the growth of four HCC cell lines at 40 µmol/l. In contrast, most of these HCC cell lines were susceptible to a low concentration of GSI-I (1.2 µmol/l) treatment. The suppressive effect of GSI-I on cell growth was because of the inhibition of C-Myc, a Notch target gene. In addition, 80% (16/20) of the specimens showed either an increased expression of at least one Notch receptor or an augmented expression of Jagged1 compared with their normal counterparts. Our study reports for the first time that different kinds of GSIs can block the growth of several HCC cell lines. Our finding suggests that GSI-I is a potential chemical reagent and warrants additional testing in liver cancer therapeutics.

Efficacy of Shaobei injection in the treatment of grade II-III hemorrhoids and the effect on fibulin protein expression: A study protocol of a randomized controlled trial.

BACKGROUND: Hemorrhoids are a common and seriously disruptive condition that seriously affects people's lives in terms of treatment. Injection therapy is an effective minimally invasive scheme for the treatment of grade II-III hemorrhoids, but its clinical application is limited by the adverse reactions caused by injection drugs. Some clinical studies have confirmed the efficacy and safety of Shaobei injection as a traditional Chinese medicine extract. However, there is no standard randomized controlled study to verify its efficacy and explore its potential mechanism. METHODS: This is a prospective, randomized, single blind, parallel controlled trial to study the efficacy of Shaobei injection in the treatment of grade II-III hemorrhoids and its effect on the expression of fibulin-3 and fibulin-5 in fibulin protein family. The patients will be randomly divided into a treatment group and control group. The treatment group will be treated with Shaobei injection, and the control group will be treated with rubber band ligation. The observation indexes include: visual analysis scale, postoperative hospital stay, total use of painkillers, fibulin-3 and fibulin-5, hemorrhoids recurrence, and adverse events. Finally, the data will be statistically analyzed by SPASS 18.0 software. DISCUSSION: This study will compare the efficacy of Shaobei injection with the rubber band ligation method in the treatment of grade II-III haemorrhoids and investigate its effect on the expression of fibulin-3 and fibulin-5 in the fibulin protein family. The results of this study will provide a basis for the clinical use of Paeoniflora injection as an alternative to traditional sclerosing agent in the treatment of grade II-III haemorrhoids.Trial registration: OSF Registration number:DOI 10.17605/OSF.IO/MKVDB.