Anticancer activity and underlying mechanism of neogambogic acid / 中国天然药物

Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese medicine with various biological functions such as detoxification, anti-inflammatory, and anthelmintic activities. Recent studies suggest that garcinia has potential anticancer activity. Increasing evidences indicate that the main active monomer gambogic acid isolated from garcinia can inhibit the growth of various cancer cells. Neogambogic acid is an isolated compound with a similar chemical structure as gambogic acid. Preliminary studies show that the neogambogic acid can selectively inhibit the growth of various cancer cells, and has a broader antitumor activity and lower toxicity than gambogic acid. In this review, we summarize the advances made in the investigation of the anti-tumor effect of neogambogic acid in recent years.

Mechanism of gambogenic acid in resisting angiogenesis of lung cancer in vitro / 中国中药杂志

The aim of this paper was to observe the effect of gambogenic acid on angiogenesis of lung cancer and its preliminary mechanism. After culturing lung adenocarcinoma A549 cells, the conditioned medium was treated with gambogenic acid and then used to culture human umbilical vein endothelial cells (HUVECs) to establish the indirect contact cell co-culture system. A two-dimensional culture model of HUVEC was established with matrigel to observe the effect of gambogenic acid on angiogenesis. DAPI staining was used to observe the morphological changes in HUVEC cells after treatment with gambogenic acid under the fluorescence microscope. Annexin V-FITC/PI staining and flow cytometry analysis were used to determine gambogenic acid's effect on HUVEC cell apoptosis rate. The protein expressions of PI3K, p-PI3K, Akt, p-Akt were measured by Western blot. PTEN-siRNA was transfected into cells, and RT-PCR was used to detect the expression levels of PI3K and Akt genes. Gambogenic acid can significantly inhibit angiogenesis, and its inhibitory effect was dose-dependent. DAPI staining showed apoptotic morphological features of HUVEC cells under fluorescence microscope. Annexin V-FITC/PI staining showed that gambogenic acid induced apoptosis in HUVECs. The results of Western blot showed that the expressions of p-PI3K and p-Akt protein were down-regulated with gambogenic acid, while the expressions of PI3K and Akt protein was insignificant. The results of RT-PCR indicated that the expressions of PI3K and Akt protein were up-regulated by PTEN siRNA. Gambogenic acid can inhibit angiogenesis in lung cancer in vitro, and the mechanism of inhibiting angiogenesis may be related to the PI3K/Akt signaling pathway.

Anticancer activity and underlying mechanism of neogambogic acid / 中国天然药物

Garcinia, a kind of dry resin secreted by Garcinia hanburyi Hook. F. G., is a traditional Chinese medicine with various biological functions such as detoxification, anti-inflammatory, and anthelmintic activities. Recent studies suggest that garcinia has potential anticancer activity. Increasing evidences indicate that the main active monomer gambogic acid isolated from garcinia can inhibit the growth of various cancer cells. Neogambogic acid is an isolated compound with a similar chemical structure as gambogic acid. Preliminary studies show that the neogambogic acid can selectively inhibit the growth of various cancer cells, and has a broader antitumor activity and lower toxicity than gambogic acid. In this review, we summarize the advances made in the investigation of the anti-tumor effect of neogambogic acid in recent years.

Photodynamic therapy with bengal rose and derivatives against leishmania amazonensis

Introduction: The treatment of cutaneous leishmaniasis [CL] is based primarily on the use of pentavalent antimonials, which may lead to many side effects limiting their use. Photodynamic therapy [PDT] is an alternative for the treatment of CL, and some xanthene dyes have the potential for use in PDT

Methods: The xanthenes rose bengal B [RB] and its derivatives rose bengal methyl ester [RBMET], and butyl ester [RBBUT] were analyzed for leishmanicidal activity against promastigotes and intracellular amastigotes of Leishmania amazonensis. Cytotoxicity was assessed in J774.A1 macrophages

Results: RB derivates RBMET [IC50 9.83 microM], and RBBUT [IC50 45.08 microM] showed leishmanicidal activity, however, were toxic to J774.A1 macrophages, resulting in low selectivity index

Conclusion: The RBMET and RBBUT showed to be effective against the L. amazonensis and the low selectivity index presented may not be a limitation for their use in PDT to CL treatment

Dormancy models for Mycobacterium tuberculosis: A minireview

Dormancy models for Mycobacterium tuberculosis play important roles in understanding various aspects of tuberculosis pathogenesis and in the testing of novel therapeutic regimens. By simulating the latent tuberculosis infection, in which the bacteria exist in a non-replicative state, the models demonstrate reduced susceptibility to antimycobacterial agents. This minireview outlines the models available for simulating latent tuberculosis both in vitro and in several animal species. Additionally, this minireview discusses the advantages and disadvantages of these models for investigating the bacterial subpopulations and susceptibilities to sterilization by various antituberculosis drugs.

Angiotensin II induced myocardial hypertrophy in neonatal rats could be attenuated by activated κ-opioid receptor via modulating the calcineurin signal pathways / 中华心血管病杂志

<p><b>OBJECTIVE</b>To observe the effect of κ-opioid receptor (κ-OR) stimulation on Angiotensin II (Ang II)-induced cardiomyocyte hypertrophy in vitro cultured myocardial cells from neonatal rats and on calcineurin (CaN) signal pathways.</p><p><b>METHODS</b>Cultured myocardial cells of neonatal rats were divided into control group, CSA (1 µmol/L) group, Ang II (1 µmol/L) group, Ang II (1 µmol/L) + U50488H (1 µmol/L) group, Ang II (1 µmol/L) + CSA (1 µmol/L) group, Ang II (1 µmol/L) + Rp-cAMPS (1 µmol/L) group, Ang II (1 µmol/L) + CSA (1 µmol/L) + U50488H (1 µmol/L) group and Ang II (1 µmol/L) + PTX5 mg/L + U50488H (1 µmol/L) group. The hypertrophic myocytes were induced by Ang II 1µmol/L before κ-OR agonist U50488H 1 µmol/L was administered. The antihypertrophic effect of κ-OR stimulation was observed in the presence of ciclosporine A (CsA) 1 µmol/L, cAMP triethyl-ammonium salt (Rp-cAMPS) 1 µmol/L, and pertussistoxin ( PTX) 5 mg/L. The total protein content was assayed by the method of Lowry. The [Ca²⁺]i was measured by confocal microscope using Fluo-3/AM as flouresecent indicator. The relative expression of CaN was determined by Western blot.</p><p><b>RESULTS</b>(1) The total protein content of Ang II group was significantly higher than that in control group (P<0.01), which could be equally reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). Total protein content of the Ang II + PTX + U50488H group and the Ang II group was similar. (2) The [Ca²⁺]i was significantly higher in Ang II group of neonatal rat cardiomyocytes than that in control group (P<0.01), which could be reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). [Ca(2+)]i was similar between the Ang II + PTX + U50488H group and the Ang II group. (3) The expression of CaN was significantly higher in Ang II group than that in control group (P<0.01), which could be significantly reduced by cotreatment with U50488H, CSA and Rp-cAMPS (P<0.01). CaN was similar between the Ang II + PTX + U50488H group and the Ang II group.</p><p><b>CONCLUSION</b>κ-opioid receptor activation could attenuate Ang II induced cardiomyocytes hypertrophy via reducing [Ca²⁺]i and downreglating CaN.</p>

Study on analgesia of oxymatrine and its relation to calcium channels / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study whether the analgesis of oxymatrine (OMT) affects N-type voltage-gated calcium channels (VGCCs).</p><p><b>METHODS</b>Totally 45 mice were randomly divided into the sham-operation group, the model group [established by partial sciatic nerve ligation (PSNL)] , and the OMT treatment group according to random digit table, 15 in each group. The dorsal root ganglions (DRG) were separated in PSNL pain model mice. Intracellular calcium concentration ([Ca2+]i) was determined with Fluo-3 AM immunofluorescent probe in cultured DRG neurons. Different protein expression levels of N-type (Cav2. 2) and L-type ( Cav1. 3) among VGCCs from brain and DRG tissues were detected with Western blot.</p><p><b>RESULTS</b>Compared with the sham-operation group, [Ca2+]i, increased in cultured DRG neurons (P <0. 05) , protein expression levels of Cav2. 2 in the brain tissue increased (P <0. 05), protein expression levels of Cav2. 2 in DRG tissues decreased in the model group (P <0. 01). Compared with the model group, [Ca2+]i, decreased in cultured DRG neurons (P < 0. 05), protein expression levels of Cav2. 2 in the brain tissue decreased (P <0. 01), protein expression levels of Cav2. 2 in DRG tissues increased in the OMT treatment group (P <0. 01). There was no statistical difference in Cav1. 3 expressions in cultured DRG neurons and the brain (P >0. 05).</p><p><b>CONCLUSION</b>Analgesic effect of OMT might be related to Cav2. 2 channel mediated calcium ion flux.</p>

In vitro antimycobacterial activity of six Cameroonian medicinal plants using microplate alamarBlue assay

Objective/background: The latest incidence of tuberculosis [TB] [per 100,000 people] in Cameroon was 243.00 as of 2011. Over the past 21 years, the value for this indicator has fluctuated between 112.00 in 1990 and 320.00 in 2003. Worldwide, this incidence has also increased, bringing back TB as a reemerging disease. On the same note, resistance to anti-TB drugs has increased, urging the search for new molecules

Methods: This study was carried out to evaluate the antimycobacterial activity of six medicinal plants on the virulent strain, H37Rv, using the microplate alamarBlue assay. Mycobacterium tuberculosis [H37Rv strain] was incubated with decreased concentrations of six plant extracts, ranging from 250 microg/mL to 31.25 microg/mL. After 7 days of incubation at 37 degree C, the effects of these plant extracts on the viability of the mycobacteria were evaluated. For each plant extract, the minimal inhibitory concentration was determined

Results: The results showed that the compounds MBC1, MBC24, MBC68, MBC81, MBC117, and MBC118 were the best candidates with minimal inhibitory concentrations of 31.25, 62.5, 125, 62.5, and 125 microg/mL, respectively

Conclusion: These results confirm and validate the traditional use of these plants to treat respiratory diseases, which could be good sources and alternatives of plant metabolites for anti-TB-drug development

Efficacy of Slimming Cream Containing 3.5% Water-Soluble Caffeine and Xanthenes for the Treatment of Cellulite: Clinical Study and Literature Review

BACKGROUND: Cellulite is a 'cottage cheese-like' cutaneous change caused by subcutaneous fat bulging into the dermis that usually leads to cosmetic problems. Slimming cream containing 3.5% water-soluble caffeine and xanthenes exhibits a lipolytic effect with penetration into the dermis. OBJECTIVE: To evaluate the efficacy and safety of slimming cream for the treatment of cellulite. METHODS: Fifteen subjects with cellulite applied slimming cream to the thighs and inner side of the upper arms twice daily for 6 weeks. Efficacy was assessed using a standard visual scale, changes in the circumferences of the thighs and upper arms, and patient satisfaction by a questionnaire at baseline, week 3, and week 6. Safety was assessed by inquiring about adverse events through questionnaires. RESULTS: The standard visual scale score improved significantly by 0.49 points (19.8%) at week 6. Thigh and upper-arm circumferences decreased by 0.7 cm (1.7%) and 0.8 cm (2.3%), respectively, at week 6. Slight itching and transient flushing were commonly reported, but no serious adverse event occurred. CONCLUSION: The slimming cream tested appears to be effective for the treatment of cellulitis without serious adverse effects. However, additional large clinical trials are required to confirm the efficacy and safety of slimming cream for the treatment of cellulitis.

Protective effect of ligustilide against glutamate-induced apoptosis in PC12 cells / 药学学报

To investigate the neuroprotective of ligustilide (LIG) against glutamate-induced apoptosis of PC12 cells, cell viability were examined by MTT assay. Flow cytometry was applied to assay cell apoptosis rate. Intracellular calcium concentration was measured by using fluorescent dye Fluo-3/AM. Cytochrome C (Cyt C), Caspase-3, Bax and Bcl-2 protein expression were assayed by western blot. The results showed that glutamate is cytotoxic with an inhibitory concentration 50 (ID50) of 15 mmol · L(-1). Pretreatment with LIG (1, 5, 15 μmol · L(-1)) significantly improved cell viability. The apoptosis rate in glutamate-induced PC12 cells was 13.39%, and decreased in the presence of LIG (1, 5, 15 μmol · L(-1)) by 9.06%, 6.48%, 3.82%, separately. Extracellular accumulation of Ca2+ induced by glutamate were significantly reduced by LIG. The results of western blot manifested that pretreatment LIG could decrease the release of Cyt C from mitochondria, down-regulate Caspase-3 protein expression and up-regulate Bcl-2/Bax ratio, thereby protects PC12 cells from apoptosis. In summary, LIG had protective effect on glutamate-induced apoptosis in PC12 cells through attenuating the increase in intracellular Ca2+ concentration, and inhibiting the release of Cyt C from mitochondria to cytoplasm.

Preparation and vitality detection of protoplast in Salvia miltiorrhiza Bunge / 生物工程学报

We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts.

Study of gambogenic acid-induced apoptosis of melanoma B16 cells through PI3K/Akt/mTOR signaling pathways / 中国中药杂志

<p><b>OBJECTIVE</b>To discuss the mechanism of gambogenic acid (GNA) in inducing the apoptosis of melanoma B16 cells.</p><p><b>METHOD</b>The inhibitory effect of GNA on the proliferation of B16 cells was measured by the methyl thiazolyl tetrazolium (MTT) assay. The effect of GNA on B16 cells was detected by the Hoechst 33258 staining. The transmission electron microscopy was used to observe the ultra-structure changes of B16 cells. The changes in PI3K, p-PI3K, Akt, p-Akt, p-mTOR, PTEN proteins were detected by the Western blotting to discuss the molecular mechanism of GNA in inducing the apoptosis of B16 cells.</p><p><b>RESULT</b>GNA showed a significant inhibitory effect in the growth and proliferation of melanoma B16 cells. The cell viability remarkably decreased with the increase of GNA concentration and the extension of the action time. The results of the Hoechst 33258 staining showed that cells processed with GNA demonstrated apparent apoptotic characteristics. Under the transmission electron microscope, B16 cells, after being treated with GNA, showed obvious morphological changes of apoptosis. The Western blot showed a time-dependent reduction in the p-PI3K and p-Akt protein expressions, with no change in p-PI3K and p-Akt protein expression quantities. The p-mTOR protein expression decreased with the extension of time, where as the PTEN protein expression showed a time-dependent increase.</p><p><b>CONCLUSION</b>GNA could inhibit the proliferation of melanoma B16 cells and induce their apoptosis within certain time and concentration ranges. Its mechanism in inducing the cell apoptosis may be related to PI3K/Akt/mTOR signaling pathways.</p>

Evaluation of some 1H-pyrazole derivatives as a dual acting antimalarial and anti-leishmanial agents

The synthesis of a novel series of 1H-pyrazole derivatives was achieved by condensation of pyrazole aldehyde 1 with hydrazine hydrate to give hydrazone 7. On the other hand, cyclization of alpha, beta-unsaturated ketone counterpart 2 using hydrazine hydrate in liquid aliphatic acids rendered compounds 4-6 and hydrazine hydrate in ethanol afforded compound 3. The later was allowed to react with aroyl chloride giving rise to compounds 8, 9. All compounds were tested for their in vivo anti-malarial and in vitro antileishmanial activities. The anti-malarial activity was performed using Plasmodium berghei infected mice, while the anti-leishmanial activity of the compounds was determined against Leishmania aethiopica promastigotes using alamar blue reduction assay. Compound 3, 1-[4-methylphenyl]-3-phenyl-4-[3-[2-thienyl]-2-pyrazolin-5-yl]-1H-pyrazole, possessed the highest anti-malarial activity with suppression of 70.26%. The highest anti-leishmanial activity was exhibited by compound 2, 1-[4-methylphenyl]-3-phenyl-4-[1-[2-thienyl]-prop-2-en-1-one]-1H-pyrazole, with an IC[50] of 0.079 microg/ml. Hydrazone 7 showed appreciable dual anti-malarial [suppression = 62.30%] and anti-leishmanial activity [IC[50] = 1.823 microg/ml]

Comparison of the in-vitro effects of bevacizumab, mitomycin-C, 5-fluorouracil,and triamcinolone acetonide on the viability of cultured human tenon's fibroblasts

@#<p style="text-align: justify;"><strong>Objective:</strong> To evaluate and compare the effects of bevacizumab, mitoinycin-C (MMC), 5-fluorouracil (5-FU), and triamcinolone acetonide (TA) on the viability of cultured human Tenon's capsule fibroblasts (cHTF) in vitro.</p> <p style="text-align: justify;"><strong>Methods:</strong> Human Tenon's fibroblasts (HTF) were harvested and cultured in a Roswell-Park-Memorial 1-Institute (RPMI) media. MMC, 5-FU, bevaciz. umab, and TA were administered to the cHTF at 3-fold decreasing concentrations starting from 20 ug, 5 mg, 25 mg, and 4 mg respectively. A negative control/untreated group containing RPMI media only was included in the study. Fibroblast cell viability was assessed using resazurin fluorim etric assay. Half¬maximal inhibitory concentration (IC50) was computed for agents which showed significant decrease in cHTF viability compared to the untreated group.</p> <p style="text-align: justify;"><strong>Results:</strong> There was no significant difference in cH IF viability between the untreated control group compared to 5-FU (p=0.97), bevacizumab (p=0.10), and TA (p=0.06) groups. Mitomycin-C showed a significant decrease in cHTF viability (p<0.001) which was dose dependent. The IC50 of MMC was computed at 12.16 ug using the prism software.</p> <p style="text-align: justify;"><strong>Conclusion:</strong> Mitomycin-C demonstrated dose-dependent decrease in viability of cultured human Tenon's fibroblasts. 5-FU, bevacizumab, and triamcinolone did not show this effect.</p> <p style="text-align: justify;"><strong>Key Words:</strong> Mitomycin-C, 5-fluorouracil, Bevaciz. umab, Tria. mcinolone acetonide, Fibroblast, Trabeculectomy</p>

Studies on effects of calycosin-7-O-β-D-glucoside on prim-O-glucosylcimifugin and cimifugin in vivo pharmacokinetics / 中国中药杂志

Study on the effects of Astragali Radix main active flavone calycosin-7-O-β-D-glucoside on Saposhnikoviae Radix main active ingredients prim-O-glucosylcimifugin and cimifugin, a UPLC-MS/MS method for simultaneous determination of prim-O-glucosylcimifugin and cimifugin in rat plasma was established, and the comparative pharmacokinetics of prim-O-glucosylcimifugin and cimifugin after oral administration of prim-O-glucosylcimifugin and calycosin-7-O-β-D-glucoside-prim-O-glucosylcimifugin to rats were carried out, which might be conductive in exploring the rationality of Astragali Radix - Saposhnikoviae Radix herb couple. Twelve male SD rats were divided into two groups. Prim-O-glucosylcimifugin and cimifugin in rat plasma of different time points after oral administration of prim-O-glucosylcimifugin and calycosin-7-O-β-D-glucoside - prim-O-glucosylcimifugin to rats were determinated. And the main pharmacokinetic parameters were investigated using DAS 3. 2. 4. The established method was rapid, accurate and sensitive for simultaneous determination of prim-O-glucosylcimifugin and cimifugin in rat plasma. The analysis was performed on a Waters Acquity BEH C18 column (2.1 mm x 100 mm, 1.7 μm) with the mixture of acetonitrile and 0.1% formic acid/water as mobile phase, and the gradient elution at a flow rate of 0.3 mL x min(-1). The analytes were detected by tandem mass spectrometry with the electrospray ionization (ESI) source and in the multiple reaction monitoring (MRM) mode. Compared with prim-O-glucosylcimifugin group, the AUC(0-t)., and AUC(0-∞) of p-O-glucosylcimifugin as well as the C(max) of cimifugin significantly increased (P < 0.05) in calycosin-7-O-β-D-glucoside-prim-O-glucosylcimifugin group. Calycosin-7-O-β-D-glucoside could enhance the absorption of prim-O-glucosylcimifugin and cimifugin and improve the bioavailability, explaining preliminarily the rationality of Astragali Radix-Saposhnikoviae Radix herb couple.

Early responses of the STAT3 pathway to platinum drugs are associated with cisplatin resistance in epithelial ovarian cancer

Cisplatin resistance remains one of the major obstacles when treating epithelial ovarian cancer. Because oxaliplatin and nedaplatin are effective against cisplatin-resistant ovarian cancer in clinical trials and signal transducer and activator of transcription 3 (STAT3) is associated with cisplatin resistance, we investigated whether overcoming cisplatin resistance by oxaliplatin and nedaplatin was associated with the STAT3 pathway in ovarian cancer. Alamar blue, clonogenic, and wound healing assays, and Western blot analysis were used to compare the effects of platinum drugs in SKOV-3 cells. At an equitoxic dose, oxaliplatin and nedaplatin exhibited similar inhibitory effects on colony-forming ability and greater inhibition on cell motility than cisplatin in ovarian cancer. Early in the time course of drug administration, cisplatin increased the expression of pSTAT3 (Tyr705), STAT3α, VEGF, survivin, and Bcl-XL, while oxaliplatin and nedaplatin exhibited the opposite effects, and upregulated pSTAT3 (Ser727) and STAT3β. The STAT3 pathway responded early to platinum drugs associated with cisplatin resistance in epithelial ovarian cancer and provided a rationale for new therapeutic strategies to reverse cisplatin resistance.

Simultaneous determination of paeoniflorin, ferulic acid, prim-O-glucosylcimifugin and 4'-O-beta-glucopyranosyl-5-O-methylvisamminol in zhengtian pills by HPLC / 中国中药杂志

To simultaneously determine paeoniflorin, ferulic acid, prim-O-glucosylcimifugin and 4'-O-beta-glucopyranosyl-5-O-methylvisamminol in Zhengtian pills. In the test, Insertil ODS-C18 column (4.6 mm x 250 mm, 5 microm) was adopted, with acetonitrile-0.05% phosphoric acid solution as the mobile phase for gradient elution. The flow rate was 1.0 mL x min(-1), the column temperature was 30 degrees C and the detection wavelength was 230 nm. According to the results of the test, paeoniflorin, ferulic acid, prim-O-glucosylcimifugin and 4'-O-beta-glucopyranosyl-5-O-methylvisamminol showed good linear relations between peak areas and sample sizes in 11.37-170.5, 2.188-32.82, 2.896-43.44, and 3.000-45.00 mg x L(-1) (r = 0.999 9, 0.999 9, 1.000 0, 1.000 0), respectively. The average recoveries (n = 6) were 102.3% (RSD 1.2%), 99.71% (RSD 1.9%), 101.2% (RSD 1.2%), and 99.40% (RSD 2.4%), respectively. The above four components were determined in five batches of samples by using the established method, with satisfactory results. The method was so simple, accurate and highly reproducible that it could be used for quality control of the four components in Zhengtian pills.

Effect of Dipyridamole on the Reactive Oxygen Species and Oxidative Stress in Trabecular Meshwork Cells

PURPOSE: To investigate the effects of dipyridamole (DPD) on the production of reactive oxygen species (ROS) and oxidative stress in cultured human trabecular meshwork cells (HTMC). METHODS: Antioxidant activity of DPD was determined by DPPH assay. Primarily cultured HTMC were exposed to 0, 20, and 50 microm DPD using serum-deprived media. The effect of DPD on the production of ROS was assessed with the DCHFDA assay. The effect of DPD on the t-butyl hydroperoxide (tBHP)-induced oxidative stress was assessed with resazurin assay. RESULTS: DPD showed significant antioxidant activity. DPD significantly decreased the production of ROS (p < 0.05) and improved cellular activity significantly after treatment with t-BHP (p < 0.05). DPD did not affect the generation of nitric oxides. CONCLUSIONS: DPD suppressed the formation of ROS and possessed cytoprotective activity against the oxidative stress in HTMC.

Different uptake of gentamicin through TRPV1 and TRPV4 channels determines cochlear hair cell vulnerability

Hair cells at the base of the cochlea appear to be more susceptible to damage by the aminoglycoside gentamicin than those at the apex. However, the mechanism of base-to-apex gradient ototoxicity by gentamicin remains to be elucidated. We report here that gentamicin caused rodent cochlear hair cell damages in a time- and dose-dependent manner. Hair cells at the basal turn were more vulnerable to gentamicin than those at the apical turn. Gentamicin-conjugated Texas Red (GTTR) uptake was predominant in basal turn hair cells in neonatal rats. Transient receptor potential vanilloid 1 (TRPV1) and 4 (TRPV4) expression was confirmed in the cuticular plate, stereocilia and hair cell body of inner hair cells and outer hair cells. The involvement of TRPV1 and TRPV4 in gentamicin trafficking of hair cells was confirmed by exogenous calcium treatment and TRPV inhibitors, including gadolinium and ruthenium red, which resulted in markedly inhibited GTTR uptake and gentamicin-induced hair cell damage in rodent and zebrafish ototoxic model systems. These results indicate that the cytotoxic vulnerability of cochlear hair cells in the basal turn to gentamicin may depend on effective uptake of the drug, which was, in part, mediated by the TRPV1 and TRPV4 proteins.

Comparative evaluation of the microplate nitrate reductase assay and the rezasurin microtitre assay for the rapid detection of multidrug resistant Mycobacterium tuberculosis clinical isolates

The microplate nitrate reductase assay (MNRA) and the rezasurin microtitre assay (REMA) were used for the susceptibility testing of 73 clinical isolates and the results were compared with those that were obtained using the Bactec 460 TB and Bactec MGIT 960 systems. The REMA and the MNRA were performed in 96-well plates. For the REMA, the concentrations of isoniazid (INH) and rifampicin (RIF) ranged from 1.0-0.01 µg/mL and 2.0-0.03 µg/mL, respectively. For the MNRA, the INH concentration was between 1.0-0.03 µg/mL and the RIF concentration was between 2.0-0.06 µg/mL. For the MNRA, the sensitivity, specificity, positive predictive value, negative predictive value and INH/RIF agreement were 100/95.6, 97.6/100, 96.8/100, 100/98 and 98.6/98.6, respectively, and for the REMA, they were 100/91.3, 90.4/100, 88.5/100, 100/96.1 and 94.5/97.2, respectively. Our data suggest that these two rapid, low-cost methods may be inexpensive, alternative assays for the rapid detection of multidrug resistant tuberculosis in low-income countries.