Neuroprotective Effects of Water Extract from Brown Algae Petalonia binghamiae in an Experimental Model of Focal Cerebral Ischemia In Vitro and In Vivo.

Focal cerebral ischemia (fCI) can result in brain injury and sensorimotor deficits. Brown algae are currently garnering scientific attention as potential therapeutic candidates for fCI. This study investigated the therapeutic effects of the hot water extract of Petalonia binghamiae (wPB), a brown alga, in in vitro and in vivo models of fCI. The neuroprotective efficacy of wPB was evaluated in an in vitro excitotoxicity model established using HT-22 cells challenged with glutamate. Afterward, C57/BL6 mice were administered wPB for 7 days (10 or 100 mg/kg, intragastric) and subjected to middle cerebral artery occlusion and reperfusion (MCAO/R) operation, which was used as an in vivo fCI model. wPB co-incubation significantly inhibited cell death, oxidative stress, and apoptosis, as well as stimulated the expression of heme oxygenase-1 (HO-1), an antioxidant enzyme, and the nuclear translocation of its upstream regulator, nuclear factor erythroid 2-related factor 2 (Nrf2) in HT-22 cells challenged with glutamate-induced excitotoxicity. Pretreatment with either dose of wPB significantly attenuated infarction volume, neuronal death, and sensorimotor deficits in an in vivo fCI model. Furthermore, the attenuation of oxidative stress and apoptosis in the ischemic lesion accompanied the wPB-associated protection. This study suggests that wPB can counteract fCI via an antioxidative effect, upregulating the Nrf2/HO-1 pathway.

Rubus fruticosus leaf extract inhibits vascular dementia-induced memory impairment and neuronal loss by attenuating neuroinflammation.

Vascular dementia (VaD) is characterized by progressive memory impairment, which is associated with microglia-mediated neuroinflammation. Polyphenol-rich natural plants, which possess anti-inflammatory activities, have attracted scientific interest worldwide. This study investigated whether Rubus fruticosus leaf extract (RFLE) can attenuate VaD. Sprague-Dawley rats were separated into five groups: SO, sham-operated and treated with vehicle; OP, operated and treated with vehicle; RFLE-L, operated and treated with low dose (30 mg/kg) of RFLE; RFLE-M, operated and treated with medium dose (60 mg/kg) of RFLE; and RFLE-H, operated and treated with high dose (90 mg/kg) of RFLE. Bilateral common carotid artery and hypotension were used as a modeling procedure, and the RFLE were intraorally administered for 5 days (preoperative 2 and postoperative 3 days). The rats then underwent memory tests including the novel object recognition, Y-maze, Barnes maze, and passive avoidance tests, and neuronal viability and neuroinflammation were quantified in their hippocampi. The results showed that the OP group exhibited VaD-associated memory deficits, neuronal death, and microglial activation in hippocampi, while the RFLE-treated groups showed significant attenuation in all above parameters. Next, using BV-2 microglial cells challenged with lipopolysaccharide (LPS), we evaluated the effects of RFLE in dynamics of proinflammatory mediators and the upstream signaling pathway. RFLE pretreatment significantly inhibited the LPS-induced release of nitric oxide, TNF-α, and IL-6 and upregulation of the MAPKs/NF-κB/iNOS pathway. Collectively, we suggest that RFLE can attenuate the histologic alterations and memory deficits accompanied by VaD, and these roles are, partly due to the attenuation of microglial activation.

Mixed Medicinal Mushroom Mycelia Attenuates Alzheimer's Disease Pathologies In Vitro and In Vivo.

Alzheimer's disease (AD) is characterized by memory impairment and existence of amyloid-ß (Aß) plaques and neuroinflammation. Due to the pivotal role of oxidative damage in AD, natural antioxidative agents, such as polyphenol-rich fungi, have garnered scientific scrutiny. Here, the aqueous extract of mixed medicinal mushroom mycelia (MMMM)-Phellinus linteus, Ganoderma lucidum, and Inonotus obliquus-cultivated on a barley medium was assessed for its anti-AD effects. Neuron-like PC12 cells, which were subjected to Zn2+, an Aß aggregator, were employed as an in vitro AD model. The cells pretreated with or without MMMM were assayed for Aß immunofluorescence, cell viability, reactive oxygen species (ROS), apoptosis, and antioxidant enzyme activity. Then, 5XFAD mice were administered with 30 mg/kg/day MMMM for 8 weeks and underwent memory function tests and histologic analyses. In vitro results demonstrated that the cells pretreated with MMMM exhibited attenuation in Aß immunofluorescence, ROS accumulation, and apoptosis, and incrementation in cell viability and antioxidant enzyme activity. In vivo results revealed that 5XFAD mice administered with MMMM showed attenuation in memory impairment and histologic deterioration such as Aß plaque accumulation and neuroinflammation. MMMM might mitigate AD-associated memory impairment and cerebral pathologies, including Aß plaque accumulation and neuroinflammation, by impeding Aß-induced neurotoxicity.

Verapamil-loaded supramolecular hydrogel patch attenuates metabolic dysfunction-associated fatty liver disease via restoration of autophagic clearance of aggregated proteins and inhibition of NLRP3.

BACKGROUND: Obesity, a serious threat to public health, is linked to chronic metabolic complications including insulin resistance, type-2 diabetes, and metabolic dysfunction-associated fatty liver disease (MAFLD). Current obesity medications are challenged by poor effectiveness, poor patient compliance, and potential side effects. Verapamil is an inhibitor of L-type calcium channels, FDA-approved for the treatment of hypertension. We previously investigated the effect of verapamil on modulating autophagy to treat obesity-associated lipotoxicity. This study aims to develop a verapamil transdermal patch and to evaluate its anti-obesity effects. METHODS: Verapamil is loaded in biomimetic vascular bundle-like carboxymethyl pullulan-based supramolecular hydrogel patches cross-linked with citric acid and glycerol linkages (CLCMP). The investigation was then carried out to determine the therapeutic effect of verapamil-loaded CLCMP (Vera@CLCMP) on diet-induced obese mice. RESULTS: Vera@CLCMP hydrogel patches with hierarchically organized and anisotropic pore structures not only improved verapamil bioavailability without modifying its chemical structure but also enhanced verapamil release through the stratum corneum barrier. Vera@CLCMP patches exhibit low toxicity and high effectiveness at delivering verapamil into the systemic circulation through the dermis in a sustained manner. Specifically, transdermal administration of this patch into diet-induced obese mice drastically improved glucose tolerance and insulin sensitivity and alleviated metabolic derangements associated with MAFLD. Furthermore, we uncovered a distinct molecular mechanism underlying the anti-obesity effects associated with the hepatic NLR family pyrin domain-containing 3 (NLRP3) inflammasome and autophagic clearance by the vera@CLCMP hydrogel patches. CONCLUSION: The current study provides promising drug delivery platforms for long-term family treatment of chronic diseases, including obesity and metabolic dysfunctions.

Red-Emitting Latex Nanoparticles by Stepwise Entrapment of ß-Diketonate Europium Complexes.

The core-shell structure of poly(St-co-MAA) nanoparticles containing ß-diketonate Eu3+ complexes were synthesized by a step-wise process. The ß-diketonate Eu3+ complexes of Eu (TFTB)2(MAA)P(Oct)3 [europium (III); 4,4,4-Trifluoro-1-(2-thienyl)-1,3-butanedione = TFTB; trioctylphosphine = (P(Oct)3); methacrylic acid = MAA] were incorporated to poly(St-co-MAA). The poly(St-co-MAA) has highly monodispersed with a size of 300 nm, and surface charges of the poly(St-co-MAA) are near to neutral. The narrow particle size distribution was due to the constant ionic strength of the polymerization medium. The activated carboxylic acid of poly(St-co-MAA) further chelated with europium complex and polymerize between acrylic groups of poly(St-co-MAA) and Eu(TFTB)2(MAA)P(Oct)3. The Em spectra of europium complexes consist of multiple bands of Em at 585, 597, 612 and 650 nm, which are assigned to 5D0→7FJ (J = 0-3) transitions of Eu3+, respectively. The maximum Em peak is at 621 nm, which indicates a strong red Em characteristic associated with the electric dipole 5D0→7F2 transition of Eu3+ complexes. The cell-specific fluorescence of Eu(TFTB)2(MAA)P(Oct)3@poly(St-co-MAA) indicated endocytosis of Eu(TFTB)2(MAA)P(Oct)3@poly(St-co-MAA). There are fewer early apoptotic, late apoptotic and necrotic cells in each sample compared with live cells, regardless of the culture period. Eu(TFTB)2(MAA)P(Oct)3@poly(St-co-MAA) synthesized in this work can be excited in the full UV range with a maximum Em at 619 nm. Moreover, these particles can substitute red luminescent organic dyes for intracellular trafficking and cellular imaging agents.

Perilla frutescens Leaf Extract Attenuates Vascular Dementia-Associated Memory Deficits, Neuronal Damages, and Microglial Activation.

Vascular dementia (VaD) is characterized by a time-dependent memory deficit and essentially combined with evidence of neuroinflammation. Thus, polyphenol-rich natural plants, which possess anti-inflammatory properties, have received much scientific attention. This study investigated whether Perilla frutescens leaf extract (PFL) exerts therapeutic efficacy against VaD. Sprague Dawley rats were divided into five groups: SO, sham-operated and vehicle treatment; OP, operated and vehicle treatment; PFL-L, operated and low-dose (30 mg/kg) PFL treatment; PFL-M, operated and medium-dose (60 mg/kg) PFL treatment; and PFL-H, operated and high-dose (90 mg/kg) PFL treatment. Two-vessel occlusion and hypovolemia (2VO/H) were employed as a surgical model of VaD, and PFL was given orally perioperatively for 23 days. The rats underwent the Y-maze, Barnes maze, and passive avoidance tests and their brains were subjected to histologic studies. The OP group showed VaD-associated memory deficits, hippocampal neuronal death, and microglial activation; however, the PFL-treated groups showed significant attenuations in all of the above parameters. Using lipopolysaccharide (LPS)-stimulated BV-2 cells, a murine microglial cell line, we measured PFL-mediated changes on the production of nitric oxide (NO), TNF-α, and IL-6, and the activities of their upstream MAP kinases (MAPKs)/NFκB/inducible NO synthase (iNOS). The LPS-induced upregulations of NO, TNF-α, and IL-6 production and MAPKs/NFκB/iNOS activities were globally and significantly reversed by 12-h pretreatment of PFL. This suggests that PFL can counteract VaD-associated structural and functional deterioration through the attenuation of neuroinflammation.

Water Extract of Mixed Mushroom Mycelia Grown on a Solid Barley Medium Is Protective against Experimental Focal Cerebral Ischemia.

Although the individual consumption of medicinal mushrooms, including Phellinus linteus (PL), Ganoderma lucidum (GL), and Inonotus obliquus (IO), is known to be neuroprotective, the associated mechanisms underlying their therapeutic synergism on focal cerebral ischemia (fCI) have yet to be elucidated. This study aimed to demonstrate the neuroprotective effects of mixed mushroom mycelia (MMM) against experimental fCI. The water-fractions, ethanolic-fractions, and ethyl acetate-fractions of the MMM (PL, GL, and IO) grown in a barley medium using solid-state fermentation techniques were prepared and their protective effects against glutamate-induced excitotoxicity were compared in PC-12 cells. After the identification of the water extracts of MMM (wMMM) as the most suitable form, which possessed the lowest toxicity and highest efficacy, further analyses for evaluating the anti-apoptotic effects of wMMM, including Hoechst 33258-based nuclear staining, fluorescence-activated cell sorting, and reactive oxygen species (ROS) detection assays, were performed. Rats were subjected to a 90 min middle cerebral artery occlusion and reperfusion, after which a wMMM treatment resulted in significant dose-dependent improvements across a number of parameters. Furthermore, measurements of intracellular ROS and levels of antioxidant enzymes revealed a wMMM-mediated ROS attenuation and antioxidant enzyme upregulation. We suggest that wMMM is neuroprotective against fCI through its anti-apoptotic and anti-oxidative effects.

Highly Luminescent and Anti-Photobleaching Core-Shell Structure of Mesoporous Silica and Phosphatidylcholine Modified Superparamagnetic Iron Oxide Nanoparticles.

Highly fluorescent magnetic nanoparticles (Eu(TTA)3(P(Oct)3)3@mSiO2@SPION) [europium (III) chloride hexahydrate = Eu; 4,4,4-trifluoro-1-(2-thienyl)-1,3-butanedione = TTA; trioctylphosphine = (P(Oct)3); mesoporous silica = mSiO2; superparamagnetic iron oxide nanoparticle = SPION] were developed as a dual-functional imaging agent. The hierarchical structure was composed of a magnetic core and mesoporous silica shell was constructed using a cationic surfactant template after coating with phosphatidylcholine of oleic acid coated SPION. Afterward, the surface and cavities of mSiO2@SPION were modified with 3-(trimethoxysilyl) propyl methacrylate (TMSPMA) as a silane coupling agent to introduce methacrylate groups. Eu(TTA)3(P(Oct)3)3 molecules are penetrated, located and bonded covalently inside of the cavities/mesopores of mSiO2, it shows extremely stable anti-photobleaching properties. The emission spectra of Eu(TTA)3(P(Oct)3)3@mSiO2@SPION indicated typical hypersensitivity transition 5D0→7F2 at 621 nm. The concentration of Eu(TTA)3(P(Oct)3)3@mSiO2@SPION was varied between 10 and 500 µL/mL to evaluate the cytotoxicity with NCI-H460 (H460) cells using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. In addition, the presence of a strong red-emitting Eu(TTA)3(P(Oct)3)3@mSiO2@SPION in the cytoplasm was observed by fluorescence microscopy. Those results that it can be a potential candidate for dual-functional contrast agent and PL nanomaterials for fabricating the diagnostic kits to amplify the low signal.

Protective Effect of Cholic Acid-Coated Poly Lactic-Co-Glycolic Acid (PLGA) Nanoparticles Loaded with Erythropoietin on Experimental Stroke.

Stroke is a major cause of adult mortality and morbidity worldwide. However, the treatment of stroke using the vast majority of possible drug candidates, including erythropoietin (EPO), remains problematic because of the presence of the blood-brain barrier (BBB), resulting in scarce penetration onto the brain. To overcome this, we synthesized a novel EPO delivery system, namely the cholic acid-coated poly lactic-co-glycolic acid (PLGA) nanoparticles loaded with EPO (EPO-CA-NPs), with the aim of enabling efficient penetration of EPO-CA-NPs across the BBB. The therapeutic efficacy of EPO-CA-NPs on an animal model of stroke was compared with that of EPO. The experimental stroke model was produced by subjecting rats to the middle carotid artery occlusion and reperfusion (MCAO/R) technique. The results indicated that EPO-CA-NPs reduced the extent of the infarct volume and cellular apoptosis to a greater extent than EPO alone at postoperative day (POD) 1. Furthermore, EPO-CA-NPs showed better performance on sensorimotor functions than EPO alone at POD 1, 3, 5, and 7. Taken together, EPO-CA-NPs, a newly synthesized brain-targeted EPO-delivery system, has stronger therapeutic effects on stroke than EPO alone, by enabling efficient EPO delivery into the brain.

Topical film prepared with Rhus verniciflua extract-loaded pullulan hydrogel for atopic dermatitis treatment.

Atopic dermatitis (AD) is characterized by relapsing pruritus and skin dryness. Due to the pathogenic multiplicity and the adverse effects associated with the current therapeutics, development of transdermal drug delivery system is becoming an area of interest. Here, a novel topical film prepared with Rhus verniciflua extract (RVE)-loaded pullulan hydrogel (RVE@PH) was synthesized and tested its therapeutic efficacy on the AD rats modeled by neonatal capsaicin injection method. The RVE@PH was characterized by a Fourier-transform infrared spectroscopy and an in vitro release assay. Rat pups were randomly divided into two groups: vehicle-treated (VEH; n = 5) and capsaicin-treated (n = 15). The latter were given capsaicin subcutaneously at 24 h after birth for AD induction and further divided into three groups (n = 5 per each): not treated (CAP), pullulan hydrogel-applied (PH), and RVE@PH-applied (RVE-PH). The pullulan hydrogel and RVE@PH were topically applied on shoulder lesions for 14 days (from 42 to 56 days after birth). Their phenotypes were compared based on the dermatitis score, epidermal thickness, mast cell infiltration, and serum myeloperoxidase (MPO) activities. The PH group showed significant attenuation in all the aforementioned values compared to the CAP group, suggesting that pullulan hydrogel itself has therapeutic activity against AD. Notably, the attenuations were more potent in the RVE-PH group than the PH group, indicating that the therapeutic efficacy against AD is augmented by the presence of RVE, a loaded pharmaceutic. Collectively, these results indicate that RVE@PH inhibits AD through exerting the dual roles, that is, the pullulan hydrogel-mediated physical and RVE-mediated pharmaceutical actions. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 2325-2334, 2019.

Magnetofluorescent Nanocomposite Comprised of Carboxymethyl Dextran Coated Superparamagnetic Iron Oxide Nanoparticles and ß-Diketon Coordinated Europium Complexes.

Red emitting europium (III) complexes Eu(TFAAN)3(P(Oct)3)3 (TFAAN = 2-(4,4,4-Trifluoroacetoacetyl)naphthalene, P(Oct)3 = trioctylphosphine) chelated on carboxymethyl dextran coated superparamagnetic iron oxide nanoparticles (CMD-SPIONs) was synthesized and the step wise synthetic process was reported. All the excitation spectra of distinctive photoluminesces were originated from f-f transition of EuIII with a strong red emission. The emission peaks are due to the hypersensitive transition 5D0→7F2 at 621 nm and 5D0→7F1 at 597 nm, 5D0→7F0 at 584 nm. No significant change in PL properties due to addition of CMD-SPIONs was observed. The cytotoxic effects of different concentrations and incubation times of Eu(TFAAN)3(P(Oct)3)3 chelated CMD-SPIONs were evaluated in HEK293T and HepG2 cells using the WST assay. The results imply that Eu(TFAAN)3(P(Oct)3)3 chelated CMD-SPIONs are not affecting the cell viability without altering the apoptosis and necrosis in the range of 10 to 240 µg/mL concentrations.

Platycarya strobilacea leaf extract protects mice brain with focal cerebral ischemia by antioxidative property.

The leaf extract of Platycarya strobilacea (PSL) has long been recognized as possessing various health-promoting activities. However, information on its possible protective effects against ischemic stroke is currently lacking. Here, using a mouse model of focal cerebral ischemia (fCI), we studied the protective potential of an oral supplement of PSL. Mice were randomly divided into four groups: SO, a group subjected to a sham-operation; VEH, pretreated with distilled water and subjected to middle cerebral artery occlusion and reperfusion (MCAO/R); PSL-L and PSL-H, pretreated with low (20 mg/kg) and high (100 mg/kg) doses of PSL, respectively, and subjected to the MCAO/R procedure. PSL was administered via an oral route daily for 8 days prior to surgery. We then measured the infarct volumes and sensorimotor deficits and studied the underlying antioxidant mechanisms by quantifying apoptosis, reactive oxygen species (ROS) generation, oxidative damages, and antioxidant enzymes in the ischemic cortex. The results showed a marked attenuation in infarct volume and sensorimotor deficits in both the PSL-L and PSL-H groups when compared with VEH. The terminal deoxynucleotidyl transferase dUTP nick end labeling and the immunohistochemical detection of the cleaved caspase-3 revealed that PSL could reduce cellular apoptosis in the ischemic lesion in a dose-dependent manner. The dihydroethidium-fluorescence, 4-hydroxynonenal, and 8-hydroxyl-2'-deoxyguanosine immunoreactivities in the ischemic lesion were markedly attenuated in the PSL-L group compared with the VEH group, indicating that PSL could attenuate ROS generation and the associated oxidative damage in the ischemic cortex. Finally, western blot results indicated that PSL can upregulate levels of heme oxygenase-1 (HO-1), an antioxidant enzyme, in the lesion area. Together, these results suggest that PSL can exert protective effects against fCI, and the mechanism may involve HO-1 upregulation.

Neuroprotective Effect of Nortriptyline in Overt Hepatic Encephalopathy Through Attenuation of Mitochondrial Dysfunction.

Hyperammonemia associated with overt hepatic encephalopathy (OHE) causes excitotoxic neuronal death through activation of the cytochrome C (CytC)-mediated mitochondria-dependent apoptotic pathway. We tested the therapeutic effect of nortriptyline (NT), a mitochondrial permeability transition pore (mPTP) blocker that can possibly inhibit mitochondrial CytC efflux to the cytosol on in vivo and in vitro OHE models. After ensuring the generation of OHE rats, established by bile duct ligation (BDL), they were intraperitoneally administered either 20 mg/kg NT (i.e., BDL+NT) or another vehicle (i.e., BDL+VEH) for 14 days. Compared with the control, BDL+VEH showed an increment of motor deficits, cell death, synaptic loss, apoptosis, and mitochondria with aberrant morphology in substantia nigra compacta dopaminergic (DA-ergic) neurons. However, the extent was significantly reversed in BDL+NT. Subsequently, we studied the neuroprotective mechanism of NT using PC-12 cells, a DA-ergic cell line, which exposed glutamate used as an excitotoxin. Compared with the control, the cells exposed to 15 mM glutamate (i.e., GLU) showed incremental cell death, apoptosis, and demise in mitochondrial respiration. Importantly, efflux of CytC from mitochondria to cytosol and the dissipation of mitochondrial membrane potential (△Ψm), an indicator of mPTP opening, were prominent in GLU. However, compared with the GLU, the cells cotreated with 10 µM NT (i.e., GLU+NT) showed a significant reduction in the aforementioned phenomenon. Together, we concluded that NT can be used for OHE therapeutics, mitigating the excitotoxic death of substantia nigra compacta DA-ergic neurons via mPTP-associated mitochondrial dysfunction inhibition.

Extract of Rhus verniciflua stokes protects against renal ischemia-reperfusion injury by enhancing Nrf2-mediated induction of antioxidant enzymes.

Ischemia-reperfusion injury (IRI) may cause acute kidney disease (AKD) by mediating the oxidative stress-induced apoptosis of parenchymal cells. The extract of Rhus verniciflua Stokes (RVS) is used as a traditional herbal medicine as it exhibits anti-oxidant, anti-apoptotic and anti-inflammatory properties. Therefore, the current study investigated the therapeutic effect and the underlying mechanism of RVS on IRI-induced AKD in vivo and in vitro. The current study assessed the effects of RVS on a mouse model of renal IRI and in hypoxic human renal tubular epithelial HK-2 cells. The results demonstrated that the IRI-induced elevation of blood urea nitrogen, serum creatinine and lactate dehydrogenase was significantly attenuated by the intraoral administration of RVS (20 mg/kg/day) for 14 days prior to surgery. It was demonstrated that IRI surgery induced histological damage and cellular apoptosis in renal parenchyma, which were attenuated by pretreatment with RVS. Furthermore, in HK-2 cells incubated with 300 µM CoCl2 to induce chemical hypoxia, it was demonstrated that RVS treatment significantly inhibited cell death and the production of reactive oxygen species (ROS). Furthermore, RVS treatment upregulated the levels of endogenous antioxidant enzymes, including heme oxygenase-1 and catalase, as well as their upstream regulator nuclear factor erythroid 2-related factor 2, in HK-2 cells. Taken together, these results suggested that the intraoral administration of RVS induces a therapeutic effect on IRI-induced AKD. These effects are at least partly due to the attenuation of ROS production via upregulation of the antioxidant defense system in renal tubular cells.

Protective effect of Rhus verniciflua Stokes extract in an experimental model of post-menopausal osteoporosis.

Post-menopausal osteoporosis (PMO) is a major global human health concern. Owing to the need for therapeutic drugs without side effects, natural extracts containing various polyphenolic compounds that may exert estrogenic effects have been studied in depth. Rhus verniciflua Stokes (RVS), which has been used as a traditional herbal medicine for centuries in Korea, was recently revealed to exert estrogenic effects attributable to its bioactive ingredients sulfuretin and butein, which have strong estrogen receptor-binding affinities. In this study, the protective potential of RVS in PMO was evaluated by using an experimental animal model of PMO, which was established by ovariectomy (OVX) of female Sprague Dawley rats. The oral administration of RVS at 20 mg/kg or 100 mg/kg for 8 weeks markedly protected against OVX-induced atrophy of the uterine tube and reversed the elevation in the ratio of serum receptor activator of nuclear factor-κB ligand to osteoprotegerin, which is a marker of disease severity. In addition, RVS inhibited OVX-induced tibia bone loss, activated osteogenic activity, and suppressed osteoclastic activity in the tibial epiphyseal plate, a region of bone remodeling. Collectively, these factors indicated that the oral intake of RVS might be beneficial for the prevention of PMO.

Intermittent fasting is neuroprotective in focal cerebral ischemia by minimizing autophagic flux disturbance and inhibiting apoptosis.

Previous studies have demonstrated that autophagy induced by caloric restriction (CR) is neuroprotective against cerebral ischemia. However, it has not been determined whether intermittent fasting (IF), a variation of CR, can exert autophagy-related neuroprotection against cerebral ischemia. Therefore, the neuroprotective effect of IF was evaluated over the course of two weeks in a rat model of focal cerebral ischemia, which was induced by middle cerebral artery occlusion and reperfusion (MCAO/R). Specifically, the role of autophagy modulation as a potential underlying mechanism for this phenomenon was investigated. It was demonstrated that IF reduced infarct volume and brain edema, improved neurobehavioral deficits, and rescued neuronal loss after MCAO/R. Furthermore, neuronal apoptosis was decreased by IF in the rat cortex. An increase in the number of autophagosomes (APs) was demonstrated in the cortices of IF-treated rats, using immunofluorescence staining and transmission electron microscopy. Using immunoblots, an IF-induced increase was detected in microtubule-associated protein 1 light chain 3 (LC3)-II, Rab7, and cathepsin D protein levels, which corroborated previous morphological studies. Notably, IF reduced the accumulation of APs and p62, demonstrating that IF attenuated the MCAO/R-induced disturbance of autophagic flux in neurons. The findings of the present study suggest that IF-induced neuroprotection in focal cerebral ischemia is due, at least in part, to the minimization of autophagic flux disturbance and inhibition of apoptosis.

Rhus verniciflua Stokes attenuates cholestatic liver cirrhosis-induced interstitial fibrosis via Smad3 down-regulation and Smad7 up-regulation.

Cholestatic liver cirrhosis (CLC) eventually proceeds to end-stage liver failure by mediating overwhelming deposition of collagen, which is produced by activated interstitial myofibroblasts. Although the beneficial effects of Rhus verniciflua Stokes (RVS) on various diseases are well-known, its therapeutic effect and possible underlying mechanism on interstitial fibrosis associated with CLC are not elucidated. This study was designed to assess the protective effects of RVS and its possible underlying mechanisms in rat models of CLC established by bile duct ligation (BDL). We demonstrated that BDL markedly elevated the serological parameters such as aspartate aminotransferase, alanine transaminase, total bilirubin, and direct bilirubin, all of which were significantly attenuated by the daily uptake of RVS (2 mg/kg/day) for 28 days (14 days before and after operation) via intragastric route. We observed that BDL drastically induced the deterioration of liver histoarchitecture and excessive deposition of extracellular matrix (ECM), both of which were significantly attenuated by RVS. In addition, we revealed that RVS inhibited BDL-induced proliferation and activation of interstitial myofibroblasts, a highly suggestive cell type for ECM production, as shown by immunohistochemical and semi-quantitative detection of α-smooth muscle actin and vimentin. Finally, we demonstrated that the anti-fibrotic effect of RVS was associated with the inactivation of Smad3, the key downstream target of a major fibrogenic cytokine, i.e., transforming growth factor ß (TGF-ß). Simultaneously, we also found that RVS reciprocally increased the expression of Smad7, a negative regulatory protein of the TGF-ß/Smad3 pathway. Taken together, these results suggested that RVS has a therapeutic effect on CLC, and these effects are, at least partly, due to the inhibition of liver fibrosis by the downregulation of Smad3 and upregulation of Smad7.

Autophagy enhancement contributes to the synergistic effect of vitamin D in temozolomide-based glioblastoma chemotherapy.

Temozolomide (TMZ), an alkylating agent, is recommended as the initial treatment for high-grade glioblastoma. TMZ is widely used, but its short half-life and the frequency of tumor resistance limit its therapeutic efficacy. In the present study, the anticancer effect of vitamin D (VD) combined with TMZ upon glioblastoma was determined, and the underlying mechanism of this effect was identified. Through cell viability, clonogenic and wound healing assays, the current study demonstrated that treatment of a C6 glioblastoma cell line with TMZ and VD resulted in significantly increased in vitro antitumor effects compared with either VD or TMZ alone. Autophagy, hypothesized to be the dominant mechanism underlying TMZ-based tumor cell death, was maximally activated in TMZ and VD co-treated C6 cells. This was demonstrated by ultrastructural observations of autophagosomes, increased size and number of microtubule-associated protein 1 light chain 3 (LC3) puncta and increased conversion of LC3-I to LC3-II. However, the extent of apoptosis was not significantly different between cells treated with TMZ and VD and those treated with TMZ alone. Addition of the autophagy inhibitor 3-methyladenine markedly inhibited the anticancer effect of TMZ and VD treatment, indicating that the chemosensitizing effect of VD in TMZ-based glioblastoma therapy is generated through enhancement of cytotoxic autophagy. TMZ and VD co-treatment also significantly inhibited tumor progression and prolonged survival duration in rat glioblastoma orthotopic xenograft models when compared with TMZ treatment alone. These in vivo results are concordant with the aforementioned in vitro results, together revealing that the combined use of TMZ and VD exerts synergistic antitumor effects on rat models of glioblastoma and may represent an effective therapeutic strategy.

Teres minor innervation and injection site / Inervación y sitio de inyección del músculo redondo menor

The aim of this study was to elucidate the injection point on the teres minor (TM) for effective injection. Thirty-two specimens from 16 adult Korean cadavers (10 males and 6 females, age ranging from 42 to 102 years) were used in the study. The reference line between the inferior point on the inferior angle of the scapula (IA) and the most prominent point on the acromial angle of the scapula (AA) on the surface were identified. The measurements expressed the two above-mentioned parameters as X and Y coordinates in relation to the reference line. The X coordinate was located at 128.1±10.4 mm (78.1±5.7 %) and the Y coordinate was located at 25.3±7.6 mm (15.5±4.8 %). The recommended site according to this can be used for injections in patients with TM stiffness.

El objetivo fue determinar el punto de inyección en el músculo redondo menor (MRM) para llevar a cabo una correcta inyección. Se utilizaron 32 muestras correspondientes a 16 cadáveres adultos coreanos (10 hombres y 6 mujeres, entre 42 a 102 años de edad). Se identificó en la superficie la línea de referencia entre el punto inferior en el ángulo inferior de la escápula (AI) y el punto más prominente en el ángulo acromial de la escápula (AA). Las mediciones expresaron los dos parámetros mencionados anteriormente como coordenadas X e Y en relación con la línea de referencia. La coordenada X se encuentra a 128,1±10,4 mm (78,1±5,7 %) y la coordenada Y estaba situada a 25,3±7,6 mm (15,5±4,8 %). El sitio recomendado de acuerdo a estos resultados se puede usar para preparaciones inyectables en pacientes con rigidez del MRM.

Median nerve location change evaluated by soft cadaver dissection according to the forearm movement / Cambio de posición del nervio mediano evaluado mediante disección cadavérica blanda según el movimiento del antebrazo

The aim of this study was to examine the change in median nerve location according to forearm movement. Thirty fresh specimens from 15 adult Korean cadavers (10 males and five females; age range, 53­91 years) were examined. We measured the motor entry point according to normal and pronated positions of the forearm. The x and y coordinates of the first motor entry point (MEP) in the normal position were 1.6±0.6 cm and 2.8±1.3 cm, respectively. The depth of the MEP was 1.5±0.3 cm. The x and y coordinates of the second MEP were 1.3±0.3 cm and 4.7±1.7 cm, respectively. The depth of the MEP was 1.4±0.2 cm. The x and y coordinates of first MEP with the forearm in the pronated position were 1.9±0.7 cm and 2.3±0.8 cm. respectively. The depth of the MEP was 1.7±0.4 cm. The x and y coordinates of the second MEP were 1.6±0.7 cm and 4.4±1.7 cm, respectively. The depth of the MEP was 1.6±0.3 cm. The differences in locations according to movement of the nerve branch were 0.3 cm, and depths were 0.2 cm. The pronated position results of this study will be clinically helpful.

El objetivo fue examinar el cambio de posición del nervio mediano de acuerdo con el movimiento del antebrazo. Se examinaron 30 muestras frescas de 15 cadáveres adultos coreanos (10 hombres y 5 mujeres) con edades entre 53­91 años. Se midió el punto de entrada motor de acuerdo con las posiciones normales y pronación del antebrazo. Las coordenadas X e Y del primer punto de entrada motor (PEM) en la posición normal fueron de 1,6±0,6 cm y 2,8±1,3 cm, respectivamente. La profundidad del PEM fue de 1,5±0,3 cm. Las coordenadas X e Y del segundo PEM fueron 1,3±0,3 cm y 4,7±1,7 cm, respectivamente. La profundidad del PEM fue 1,4±0,2 cm. Las coordenadas X e Y del primer PEM del antebrazo en posición de pronación fueron 1,9±0,7 cm y 2,3±0,8 cm, respectivamente. La profundidad del PEM fue 1,7±0,4 cm. Las coordenadas X e Y del segundo PEM fueron 1,6±0,7 cm y 4,4±1,7 cm, respectivamente. La profundidad del PEM fue 1,6±0,3 cm. Las diferencias en las ubicaciones de acuerdo con el movimiento del ramo nervioso fueron de 0,3 cm, y las profundidades fueron de 0,2 cm. Los resultados de la posición de pronación de este estudio serán útiles en trabajos clínicos.