An 83-year-old woman taking isoniazid (INH) suddenly developed a headache, palpitations, and skin eruptions with itching while eating raw tuna. The symptoms were compatible with those of histamine intoxication. When given fresh raw tuna to eat, no such reactions were evoked in this patient. The episode was speculated to be due to two conditions; eating spoiled raw tuna and at the same time being on a course of INH, a potent histaminase inhibitor. We should therefore be aware of the possible adverse effects due to the interactions between drugs and foods.

The mutagenic potency of isoniazid (INH) (a widely used antitubercular drug) towards Salmonella typhimurium strain hisG46 was studied in the Salmonella/hepatocyte suspension-assay, comprising isolated human or rat hepatocytes as metabolic system. The potency of INH to induce DNA-excision repair in these hepatocytes was also measured. With rat hepatocytes, INH appeared to be only weakly mutagenic and did not induce significant increases in hepatocellular DNA-excision repair. With isolated hepatocytes of two human subjects, INH appeared also only weakly mutagenic. However, with hepatocytes of two other human subjects, INH was found to be highly mutagenic. Comparable results were obtained for the induction of hepatocellular DNA-excision repair.

BACKGROUND

Because of current limitations in improving metabolic support to the brain during hypothermic circulatory arrest (HCA), attenuation of ischemia-reperfusion injury remains an area of therapeutic intervention of relevance. Apotransferrin (Apo-Tf), alpha 1-acid glycoprotein (AGP), and C1-esterase inhibitor (C1-INH) have been herein evaluated as potential beneficial agents in reducing the ischemia-reperfusion injury in a surviving model of HCA.

METHODS

Apo-Tf 100 mg/kg (n = 6), C1-INH 50 IU/kg (n = 6), AGP 100 mg/kg (n = 6), or NaCl 0.9% 2 ml/kg (n = 6) were randomly administered to 24 juvenile pigs after a 75-min period HCA at a brain temperature of 18 degrees C.

RESULTS

Animals in the Apo-Tf group had a slightly better 7-day survival (66.7%) compared with the other study groups (50%), but such a difference was not statistically significant. Some favorable changes in the brain glucose metabolism parameters were observed in the AGP, C1-INH, and Apo-Tf groups, but these did not reach statistical significance. Semiquantitative analysis of the histopathological findings did not show any significant difference between the study groups. However, only two out of four surviving animals in the Apo-Tf group developed brain infarction, whereas all three survivors of the remaining study groups developed brain infarction.

CONCLUSIONS

Although the small size of the study groups may affect the present findings, none of the metabolic and hemodynamic parameters as well as outcome endpoints indicate a substantial therapeutic efficacy of Apo-Tf, AGP, and C1-INH as neuroprotective agents after experimental HCA.

Liver injury is a major hindrance to the treatment of tuberculosis (TB) patients due to the primary side effects associated with anti-TB drugs. Several investigations have identified sensitive biomarkers for the early diagnosis of anti-TB drug-induced liver injury (ADLI), including the use of microRNAs (miRNAs/miRs). However, the association between miR-122/155 and ADLI remains unknown. Thus, the present study used reverse transcription-quantitative polymerase chain reaction to observe changes in tissue miR-122/155 expression levels during the course of liver injury in mice. Liver injury was induced by the administration of isoniazid (INH), a first-line anti-TB drug. miR-122/155 expression levels were quantified at seven time points throughout 1 day (0.25, 0.75, 1.5, 6, 12, 18 and 24 h) based on the pharmacokinetics of INH in mice. Notably, over the timecourse of INH-induced liver injury, the tissue miR-122 expression level significantly decreased at 0.25 h, which is the peak concentration time of INH, compared with the control group (P<0.05). The change was more rapid than that of the serum aminotransferase and miR-155, which were significantly increased at 0.75 h. In addition, the pathological score correlated with the ratio of miR-122/miR-155 (r=-0.779; P<0.01). In conclusion, the miR-122/155 ratio may be utilized as a sensitive biomarker for ADLI, which could contribute to the early diagnosis of patients following anti-TB treatment.

In this work, effect of phentolamine as antagonist of alpha-1 adrenergic receptor in patients with bronchial asthma and with obstructive chronic bronchitis was studied. Parameters of the lung function are determined by body plethysmography. Raw and ITGV were registered and SRaw was calculated as well. Aerosolization is done with standard aerosolizing machines--Asthma with a possibility of aerosolization of 0.5 ml per minute. Results gained by this research shows that blockage of alpha-1 adrenergic receptor with phentolamine (10 mg by inhalator and intravenous ways and 20 mg by inhalator ways) has not changed significantly (p>> 0.1) the bronchomotor tonus of tracheobronchial tree, by comparing it with the inhalation of natrium chloride solution with percentage of 0.9% (p>> 0.1), or of hexoprenaline (2 inh x 0.2 mg) and atropine 1 mg/ml (p < 0.01). This suggests that the activity of alpha-1 adrenergic receptor in the smooth musculature is not a primary mechanism that causes reaction in patients with increased bronchial reactibility, in comparison to agonists of beta2--adrenergic receptor and cholinergic antagonists that expresses their significant action in reduction of specific resistance of airways.

Quantitative in vitro studies of antioxidant activities have been performed under aerobic conditions. However, since the biological system has lower oxygen tension, the effectiveness of antioxidants may be considerably different in vivo. alpha-Tocopherol, in vivo the most active tocopherol, is a very poor antioxidant in vitro. To clarify these points, the radical-scavenging activities of vitamin E (Toc) (alpha-, beta-, gamma- and delta-tocopherols) and ubiquinone were evaluated by the induction period method from the kinetics of polymerization of methyl methacrylate (MMA) initiated by thermal decomposition of 2,2'azobisisobutyronitrile (AIBN) (alkyl radical, R*), or benzoyl peroxide (BPO) (peroxyl radical, PhCOO*) under nearly anaerobic conditions. The ratio of the rate constant of inhibition to that of propagation (k(inh)/k(p)) for Toc was about 10 in a system with a molar ratio of AIBN to Toc of 100:1, whereas in the corresponding BPO system k(inh)/k(p) declined in the order alpha (47)>> beta (15)>> gamma (10)>> delta (7). In contrast, with AIBN the number of free radicals trapped by the phenolic moiety (n) declined in the order delta (3.0)>> gamma (2.5)>> alpha (2.2)>> beta (1.6), whereas with BPO n declined in the order delta (1.9)>> gamma (1.4)>> beta (1.0)>> alpha (0.3). A similar tendency was found in systems with a molar ratio of 10:1. Also, ubiquinone-10 showed radical-scavenging activity, although the n (0.02) was much less than that for Toc. The low n value for alpha-Toc (n = 0.3) may be attributed to the formation of stable alpha-Toc during the induction period. With a n = about 1 for beta- and gamma-Toc, a dimerization coupling of Tocs is suggested. Thus, the radical-scavenging activity is affected by the number and position of the methyl groups in the benzene nucleus of the various tocopherol compounds.

We have previously produced transgenic (TG) mice expressing the mouse inhibin alpha-subunit promoter/Simian virus 40 T-antigen (Inhalpha/Tag) fusion gene. The mice develop gonadal somatic cell tumors at the age of 5-7 months; the ovarian tumors originate from granulosa cells, and those of the testes from Leydig cells. In the present study another TG mouse line was produced, expressing under the same inh-alpha promoter the herpes simplex virus thymidine kinase gene (Inhalpha/TK). Crossbreeding of the two TG mouse lines resulted in double TG mice (Inhalpha/TK-Inhalpha/Tag), which also developed gonadal tumors. The single (Inhalpha/Tag) and double TG (Inhalpha/TK-Inhalpha/Tag) mice, both bearing gonadal tumors, were treated at the age of 5.5-6.5 months with ganciclovir (GCV, 150 mg/kg body weight twice daily i.p.) for 14 days, or with aciclovir (ACV, 300-400 mg/kg body weight per day perorally) for 2 months. During GCV treatment, the total gonadal volume including the tumor, decreased in double TG mice by an average of 40% (P<0.05), while in single TG mice, there was a concomitant increase of 60% in gonadal size (P<0.05). GCV was also found to increase apoptosis in gonads of the double TG mice. Peroral treatment with ACV was less effective, it did not reduce significantly the gonadal volume. We also analyzed the in vitro efficacy of ACV and GCV treatments in transiently HSV-TK-transfected KK-1 murine granulosa tumor cells, originating from a single-positive Inhalpha/Tag mouse. GCV proved to be more effective and more specific than ACV in action. These results prove the principle that targeted expression of the HSV-TK gene in gonadal somatic cell tumors is potentially useful for tumor ablation by antiherpes treatment. The findings provide a lead for further development of somatic gene therapy for gonadal tumors.

Tuberculosis remains as a major public health risk which causes the highest mortality rate globally and an improved regimen is required to treat the drug-resistant strains. Pyrazinamide is a first-line antitubercular drug used in combination therapy with other anti-TB drugs. Herein, we describe the modification of pyrazinamide structure using bioisosterism and rational approaches by incorporating the 1,2,3-triazole moiety. Three sets of pyrazine-1,2,3-triazoles (3a-o, 5a-o and 9a-l) are designed, synthesized and evaluated for their in vitro inhibitory potency against mycobacterium tuberculosis H₃₇Rv. The pyrazine-1,2,3-triazoles synthesized through the bioisosteric modification displayed improved activity as compared to rationally modified pyrazine-1,2,3-triazoles. Among 42 title compounds, seven derivatives demonstrated significant anti-tubercular activity with the MIC of 1.56 μg/mL, which are two-fold more potent than the parent compound pyrazinamide. Further, the synthesized pyrazinamide analogs demonstrated moderate inhibition activity against several bacterial strains and possessed an acceptable in vitro cytotoxicity profile as well. Additionally, the activity profile of pyrazine-1,2,3-triazoles was validated by performing the molecular docking studies against the Inh A enzyme. Furthermore, in silico ADME prediction revealed good oral bioavailability for the potent molecules.

The first component of human complement was separated from C1-INH by sucrose linear gradient ultracentrifugation. Activation of C1 was studied in the absence and presence of immune complexes; activation was monitored by SDS-PAGE and Western blot. When the partially purified native C1 preparation was incubated at 37 degrees C without immune complexes, activated C1s appeared after 30 min in the case of eightfold dilution with respect to the original serum, and after 45 min with 32-fold dilution. Kinetics of appearance of activated C1r was the same as that of activated C1s. From the following results, we concluded that spontaneous activation may be partially due to proteolytic enzymes contaminating the preparation: 1) a nonspecific protease inhibitor, PMSF, completely inhibited spontaneous activation but did not inhibit the activation of C1 by immune complexes; 2) alpha 2-macroglobulin partially inhibited spontaneous activation, and 3) although spontaneous activation in the absence of PMSF was relatively slow, activated C1 accelerated spontaneous activation that was completely blocked by C1-INH. In contrast to spontaneous activation, the partially purified native C1 was rapidly activated by immune complexes: within 5 min almost all C1 was activated by rabbit IgG anti-human IgM-human IgM complexes. These results support conclusions derived from activation studies when using native C1 and hemolytic assays, and do not support those derived from the activation studies with reconstituted C1 and SDS-PAGE analysis. We suggest that the contradictions can be resolved if one assumes that C1 activation can be both an intra- and intermolecular process; which process dominates is determined by the state of C1 and by experimental conditions.

OBJECTIVE

Free tissue transfer is a powerful reconstructive surgical technique. The ischemia reperfusion injury (IRI) at revascularization affects the flap and the patient; reducing this insult could improve outcomes. This study evaluated the effect of C1 esterase inhibitor (C1-inh) on IRI in a porcine musculocutaneous flap model.

METHODS

A musculocutaneous flap was transferred from the limb to the neck of 12 swine. Flaps underwent a 3-hour ischemic interval prior to revascularization. Intervention group flaps (n = 6) were perfused intra-arterially with 100U C1-inh at the commencement of the ischemic period; controls (n = 6) received heparinized saline solution. Protocol duration was 14 days; markers of reperfusion injury (creatine kinase [CK], aspartate transaminase [AST], tumor necrosis factor-alpha) were evaluated.

RESULTS

All flaps from the intervention group were viable at 14 days; five of six control flaps were viable at 14 days (P = 1). Systemic levels of biomarkers of tissue necrosis and inflammation were reduced in the intervention group. On post-operative day one, statistically significant reductions in mean levels of AST and CK were demonstrated (2,293 ± 1 × 103 U/L vs. 1,586 ± 767 U/L [P = 0.04] and 429 × 103 ± 214 × 103 U/L vs. 213 × 103 ± 156 × 103 U/L [P = 0.002], respectively). Flaps of both groups healed in their recipient locations, no adverse reactions were observed.

CONCLUSIONS

C1-inh is protective of IRI and may have utility in free tissue transfer, vascularized composite allotransplantation, and spare parts surgery. © 2016 Crown copyright. Microsurgery © 2016 Wiley Periodicals, Inc. Microsurgery 37:142-147, 2017.

The objective of this study was to determine whether neutralizing endogenous inhibin would affect ovulation rate and serum concentrations of FSH, LH, estradiol-17 beta, and progesterone in gilts. At wk 0, during their second postpubertal estrous cycle, gilts (195 +/- 2.4 d of age) were given a primary immunization against the 1-26 gly-tyr NH-terminal amino acid sequence of bovine inhibin-alpha conjugated to human alpha globulin (INH; n = 10) or against human alpha globulin alone (control; n = 10). The primary immunization mixed with Freund's complete adjuvant contained .915 mg of the inhibin peptide. Booster immunizations in Freund's incomplete adjuvant contained .3 and .183 mg of the inhibin peptide and were given at wk 8 and 12, respectively. Free, unconjugated inhibin was given to INH gilts at 16 wk. Blood samples for determination of hormones were collected every 4 h beginning on d 15 of the first estrous cycle beyond wk 16 (first cycle) and continuing until d 5 of the second estrous cycle following wk 16 (second cycle). Ovulation rate was estimated by laparoscopy during the second cycle. Antibody titers were estimated by determining the percentage of [125I]-INH bound by serum diluted 1:4,000. The antibody titers were 17 +/- 2, 22 +/- 3, and 9 +/- 1% at wk 9, 17, and 23 for INH gilts, respectively, and 0% at all times for control gilts. Duration of three consecutive estrous cycles terminating with the first experimental cycle did not differ between treatments (INH, 20.7 +/- .3 vs control, 20.4 +/- .3 d).(ABSTRACT TRUNCATED AT 250 WORDS)

BACKGROUND

Recognizing a potential interaction between isoniazid (INH), a weak monoamine oxidase inhibitor, and serotonin reuptake inhibitors (SSRIs), we assessed medication discontinuation rates in human immunodeficiency virus-infected individuals taking an SSRI, INH, or both.

METHODS

We retrospectively reviewed treatment records to determine if patients on an SSRI, INH, or both completed drug therapy in accordance with a treatment plan (e.g., 12 months of INH therapy). Patients on both medications constituted the study group; patients taking either an SSRI or INH alone constituted comparison groups.

RESULTS

There were no significant differences between the groups based on age, gender, CD4%, or CD4 count. Seven of the 10 patients (70%) in the study group discontinued therapy, which was significantly greater than the 2 of 14 (14%) in the SSRI group (P = 0.01) and the 4 of 18 (22%) in the INH group (p = 0.02) who discontinued therapy.

CONCLUSIONS

Medication discontinuation rates for patients prescribed an SSRI coincident with INH were significantly higher than for individuals prescribed these medications separately. These differences cannot be accounted for on the basis of age, gender, or CD4%, but they may be attributable to increased side effects caused by interactions between these medications.

In higher primates, increased circulating follicle-stimulating hormone (FSH) levels seen during late menstrual cycle and during menstruation has been suggested to be necessary for initiation of follicular growth, recruitment of follicles and eventually culminating in ovulation of a single follicle. With a view to establish the dynamics of circulating FSH secretion with that of inhibin A (INH A) and progesterone (P(4)) secretions during the menstrual cycle, blood was collected daily from bonnet monkeys beginning day 1 of the menstrual cycle up to 35 days. Serum INH A levels were low during early follicular phase, increased significantly coinciding with the mid cycle luteinizing hormone (LH) surge to reach maximal levels during the mid luteal phase before declining at the late luteal phase, essentially paralleling the pattern of P(4) secretion seen throughout the luteal phase. Circulating FSH levels were low during early and mid luteal phases, but progressively increased during the late luteal phase and remained high for few days after the onset of menses. In another experiment, lutectomy performed during the mid luteal phase resulted in significant decrease in INH A concentration within 2 hr (58.3+/-2 vs. 27.3+/-3 pg/mL), and a 2- to 3-fold rise in circulating FSH levels by 24 hr (0.20+/-0.02 vs. 0.53+/-0.14 ng/mL) that remained high until 48 hr postlutectomy. Systemic administration of Cetrorelix (150 microg/kg body weight), a gonadotropin releasing hormone receptor antagonist, at mid luteal phase in monkeys led to suppression of serum INH A and P(4) concentrations 24 hr post treatment, but circulating FSH levels did not change. Administration of exogenous LH, but not FSH, significantly increased INH A concentration. The results taken together suggest a tight coupling between LH and INH A secretion and that INH A is largely responsible for maintenance of low FSH concentration seen during the luteal phase.

We have studied the effect of isonicotinic acid hydrazide (INH), a convulsant agent, on the extracellular levels of amino acids in the hippocampus, and the effect of sodium valproate (VPA) administration in INH-treated rats. INH (250 mg/kg) caused a rapid and sustained decrease in basal levels of GABA, and during this period convulsions of increasing severity were observed. Basal levels of glutamine, taurine, aspartate, and glutamate were unchanged by INH. When VPA was coadministered with INH, basal GABA levels were increased and no convulsions were observed. When transmitter release was evoked using 100 mM K+, the increase in dialysate GABA observed in INH-treated animals was less than that seen in controls and convulsions increased in frequency. K(+)-evoked release of glutamate and aspartate tended to be higher following INH treatment, and in the case of aspartate, this increase was significant. VPA reversed the changes in evoked release of glutamate and aspartate, and release of GABA was considerably greater than that seen in control or INH-treated rats. No drug effect on evoked changes in taurine or glutamine level was seen. These are the first data to show decreased extracellular GABA in conjunction with convulsions in freely moving animals in vivo.

Tuberculostatic drugs are the most common drug groups with global hepatotoxicity. Awareness of potentially severe hepatotoxic reactions is vital, as hepatic impairment can be a devastating and often fatal condition. The treatment problems that may arise, within this class of medicines, are mainly of two types: adverse reactions (collateral, toxic or hypersensitive reactions) and the initial or acquired resistance of Mycobacterium tuberculosis to one or more antituberculosis drugs. Prevention of adverse reactions, increase treatment adherence and success rates, providing better control of tuberculosis (TB). In this regard, obtaining new drugs with low toxicity and high tuberculostatic potential is essential. Thus, in this work, we have designed or synthesized new derivatives of isoniazid (INH), such as new Isonicotinoylhydrazone (INH-a, INH-b and INH-c). These derivatives demonstrated good biocompatibility, antimicrobial property similar to that of parent isoniazid and last but not least, a significantly improved Pharmacotoxicological profile compared to that of isoniazid.

The objective of the present study was to evaluate the effects of novel DNA vaccines fusing inhibin (INH) α (1-32) and RFamide-related peptide-3 (RFRP-3) genes on the immune response, reproductive hormone levels, and fertility of Tan sheep. Thirty-two female Tan sheep were divided into four groups (groups A, B, C, and D with 8 sheep per group) and respectively immunized (thrice, 20 d apart) with 0.6mg of p-TPA-SINH/TPA-SRFRP (group A), p-SINH/SRFRP (group B), p-SINH (group C) or 0.4ml saline (group D). Twenty days after primary immunization, all vaccines elicited significant immune responses, and the antibody levels of anti-INH and anti-RFRP-3 in the vaccinated groups were significantly higher (p<0.05) than that of the control group. Immunization with p-TPA-SINH/TPA-SRFRP induced higher antibodies against INH and RFRP-3. Hormone levels of FSH and LH in group A immunized with p-TPA-SINH/TPA-SRFRP were significantly higher (p<0.05) than those in group C, which are immunized with p-SINH and the control group 20 d after the third immunization. Additionally, the p-TPA-SINH/TPA-SRFRP, p-SINH/SRFRP, p-SINH and saline vaccine induced different twinning rate of ewes (37.5%, 37.5%, 12.5%, and 0, respectively), but no significant differences were found in improving twinning rate of ewes among four groups. These results suggested that neutralization of endogenous INH and RFRP-3 with novel DNA vaccine fusing INH α (1-32) and RFRP-3 genes successfully elicited a humoral immune response, increased reproductive hormone levels, but it did not significantly improve litter sizes and twinning rate of ewes in the present study.

C1-inhibitor (C1-INH) is an alpha-2-neuraminoglycoprotein with a molecular weight of 105,000 daltons. It has a broad spectrum control of the many blood cascades including the complement system. Inherited deficiency of this molecule has been associated with the development of hereditary angioneurotic edema (HANE), a dominant genetic disorder. The liver and monocytes are the primary sites of C1-INH synthesis. The genetic basis of dysfunctional C1-INH is a defect at the structural locus for one C1-INH gene; both the dysfunctional C1-INH gene and the normal C1-INH gene products are present in the plasma of the affected person. The absence of C1-INH permits spontaneous activation of the first component of the complement system (C1); this, in turn, activates C4 and C2. A kinin derived from C2 may be elaborated, which then increases vascular permeability. However, recent investigations have indicated that kinin activity generated from C1-INH-depleted plasma is not derived from C2 but implicates kallikrein, an enzyme which is also controlled by C1-INH, in the formation of a kinin which is most probably bradykinin. A number of therapeutic approaches have been used to treat HANE patients, including antifribrinolytic drugs such as tranexamic acid, plasma infusion, and steroids. The anabolic steroids such as danazol and stanazolol have been used widely to treat HANE patients. We discuss in this review the potential mechanisms by which danazol promotes selective synthesis of C1-INH and several other proteins in the liver.

To examine whether there are age differences in agonist-mediated alpha 2-adrenergic receptor (alpha 2-AR) regulation, we studied the effect of a sustained increase in plasma norepinephrine (pNE) during a 7-day 10-meq Na+/day diet on platelet alpha 2-AR binding and its linked adenylate cyclase (AC) activity in 11 elderly and 11 young healthy subjects. In the young, a 41% increase in mean pNE after a low-sodium diet was correlated with a decline in receptor density (Bmax; r = -0.816; P less than 0.01) and was accompanied by a reduction in the maximal percent inhibition of sodium fluoride-stimulated AC activity by epinephrine (%AC INH; 33 +/- 4 vs. 24 +/- 4%, mean +/- SE; P less than 0.05 vs. normal diet). Despite a comparable 39% increase in mean pNE in the elderly, neither Bmax nor %AC INH was significantly reduced after a low-sodium diet. The amount of pertussis toxin substrate (Gi protein) was similar in both groups before and after dietary sodium restriction. At comparable pNE, %AC INH in the groups was similar (young, 24 +/- 4 vs. elderly, 18 +/- 4%; P = NS). We postulate that higher basal pNE levels in the elderly on normal diet may account for the lack of further downregulation of platelet alpha 2-AR density and response after low-sodium diet.

Fractionation of mouse serum by precipitation with a critical amount of polyethylene glycol 6000 (PEG; 11% w/v) results in a classical and alternative pathway-independent activation of the terminal complement route. The activation can take place after the separation of an activating principle together with the terminal route components from a natural regulator. The isolation and identification of the regulatory component preventing this activation in serum, is subject of this paper. The regulator was purified by fractionated PEG-precipitation (15-25%), followed by heparin-Sepharose affinity, Mono Q anion-exchange, and Superose 12 gel filtration chromatography. The regulator appeared to be a single-chain protein with a Mr of 96 k. A protein with similar activity purified from human serum had a Mr of 104 k and was functionally and antigenically indistinguishable from C1-INH. The mouse 96 k protein inhibited C1-esterase activity indicating that this protein is indeed C1-INH. Mouse C1-INH regulates the PEG fractionation-induced bypass activation of complement, but does not interfere with the assembly or the lytic activity of membrane attack complexes. alpha 2-Macroglobulin appeared also to be capable of inhibiting the PEG-precipitation-induced activation process, but with lower efficiency.

The effect of the phytoestrogen, quercetin (QT) on the reproductive toxicity of atrazine (ATZ) was explored in interstitial Leydig cells (ILCs). We measured the mRNA expressions of steroidogenesis genes: steroidogenic acute regulatory protein (StAR), 3β-hydroxysteroid dehydrogenase (3β-HSD), cytochrome-P450 (CYP) 11A1, insulin-like factor 3 (INSL-3), CYP17A1, inhibin-α (INH-α), androgen receptor (AR), estrogen receptor-α (ER-α), and luteinising hormone receptor (LHR) in isolated ILCs by real-time-PCR after cultured cells were treated in vitro with ATZ (232 μM) and QT (50 μM). The mRNA expression of tested genes increased with ATZ treatment and was normalized by quercetin except AR and ER-α expression. Treatment of cells with QT alone (15-50 μM) caused a dose-dependent increase in AR and ER-α mRNA expression. We also found that QT (50 μM) increased the expressions of AR and ER-α in the presence of a sub-threshold level of cyclic-AMP at 1h culture period to the levels seen with maximal stimulation of cyclic-AMP. Furthermore, the expressions of tested genes were unaffected by cyclic-AMP at 6h when the stimulatory effects of ATZ on tested genes were sustained. These findings suggest that ATZ may stimulate the expression of tested steroidogenesis genes in ILCs via a mechanism independent of cyclic-AMP which was partially antagonize by QT.

We have developed a transgenic (TG) mouse model for tumorigenesis of gonadal somatic cells using a 6 kb fragment of the mouse inhibin-alpha subunit promoter (Inh-alpha) fused with the simian virus 40 T-antigen (Tag) coding sequence. Gonadal tumors, of Leydig or granulosa cell origin, develop in the TG mice with 100% penetrance by the age of 5-8 months. Conspicuously, if the mice are gonadectomized, they develop adrenal tumors. Gonadal and adrenal tumorigenesis in these mice seem to be gonadotropin dependent. On the other hand, testosterone stimulates the proliferation of a cell line (C alpha 1) established from one of the adrenal tumors. The purpose of the present study was therefore to investigate further whether testosterone affects the growth of these gonadal and adrenal tumors in vivo. Two experimental models were used: (1) Tag TG/hypogonadotropic (hpg) double mutant mice and (2) castrated Tag TG mice. Both were treated between 1-2 and 7-8 months of age with Silastic rods (length 2 cm) containing testosterone. None of the control or testosterone-treated Tag/hpg mice developed gonadal or adrenal tumors. The castrated Tag TG mice displayed, upon microscopical examination, early stages of adrenal tumors, whereas those receiving testosterone did not show such changes. Testosterone increased the weights of gonads in the Tag/hpg mice, and those of uteri and seminal vesicles in both groups. In contrast, the adrenal weights were significantly reduced in both groups by testosterone treatment. Gonadal histology of the testosterone-treated mice showed hyperplasia of testicular Leydig cells and ovarian stroma. Spermatogenesis was induced by testosterone in the Tag/hpg mice. Adrenal histology of the testosterone-treated animals demonstrated the disappearance of the X-zone. Serum levels of FSH in testosterone-treated Tag/hpg mice were significantly increased, while those of serum LH were decreased. In conclusion, the present result indicate that the suppression of gonadotropins by testosterone implants in castrated Inh-alpha/Tag TG mice prevents the tumorigenesis of their adrenals. In intact Tag/hpg mice, testosterone implants were not able to induce gonadal or adrenal tumorigenesis. Although testosterone treatment was able to induce interstitial cell hyperplasia in gonads of the Inh-alpha/Tag mice, direct gonadotropin action is responsible for gonadal and adrenal tumorigenesis.

BACKGROUND

Vaginal lubrication, an indicator of sexual arousal and tissue health, increases significantly during genital sexual arousal. Adrenergic alpha-receptors (AR) are an important regulator of genital physiological responses involved in mediating vascular and nonvascular smooth muscle contractility; the role of β-AR in sexual arousal, however, has not yet been investigated.

OBJECTIVE

The goal of this study was to reveal the functional role of β-AR in modulating vaginal lubrication during sexual arousal and the mechanisms underlying the process.

METHODS

The effects of adrenaline on vaginal epithelial ion transport, intracellular cyclic adenosine monophosphate (cAMP) content ([cAMP]i ), and vaginal lubrication were investigated using short-circuit current (ISC ) of rat vaginas incubated in vitro, enzyme-linked immunosorbent assay (ELISA), and measurement of vaginal lubrication in vivo, respectively. The expressions of β-AR in vaginal epithelium were analyzed by reverse transcription-polymerase chain reaction, western blot, and immunofluorescence.

METHODS

Changes of ISC responses; mRNA, protein expressions and localization of β-AR; [cAMP]i ; vaginal lubrication.

RESULTS

Serosal application of adrenaline induced an increase of ISC across rat vaginal epithelium that blocked by propranolol, a β-AR antagonist, rather than phentolamine, an α-AR antagonist. β1/2-AR were both present in rat and human vaginal epithelial cells. Removing Cl(-) or application of CFTR(inh) -172, an inhibitor of cystic fibrosis transmembrane conductance regulator (CFTR), abolished adrenaline-induced ISC responses. The elevated levels of [cAMP]i induced by adrenaline were prevented by the pretreatment with propranolol. Vaginal lubrication measured in vivo showed that adrenaline or pelvic nerve stimulation caused a marked increase in vaginal lubrication, whereas pretreatment with propranolol or CFTR(inh) -172 reduced the effect.

CONCLUSIONS

Activation of epithelial β-AR facilitates vaginal lubrication during sexual arousal by stimulating vaginal epithelial Cl(-) secretion in a cAMP-dependent pathway. Thus, vaginal epithelial β-AR might be another regulator of vaginal sexual arousal responses.