Factors Predicting Prolonged Mechanical Ventilation in Guillain-Barré Syndrome.

BACKGROUND: Up to 30% of patients with Guillain-Barré syndrome require mechanical ventilation and 5% die due to acute complications of mechanical ventilation. There is a considerable group of patients that will need prolonged mechanical ventilation (considered as >14 days) and should be considered for early tracheostomy. The objective of this study is to identify risk factors for prolonged mechanical ventilation. METHODS: We prospectively analyzed patients with Guillain-Barré diagnosis with versus without prolonged mechanical ventilation. We considered clinical and electrophysiological characteristics and analyzed factors associated with prolonged mechanical ventilation. RESULTS: Three hundred and three patients were included; 29% required mechanical ventilation. When comparing the groups, patients with prolonged invasive mechanical ventilation (IMV) have a lower score on the Medical Research Council score (19.5 ± 16.2 vs 27.4 ± 17.5, p = 0.03) and a higher frequency of dysautonomia (42.3% vs 19.4%, p = 0.037), as well as lower amplitudes of the distal compound muscle action potential (CMAP) of the median nerve [0.37 (RIQ 0.07-2.25) vs. 3.9 (RIQ1.2-6.4), p = <0.001] and ulnar nerve [0.37 (RIQ0.0-3.72) vs 1.5 (RIQ0.3-6.6), p = <0.001], and higher frequency of severe axonal damage in these nerves (distal CMAP ≤ 1.0 mV). Through binary logistic regression, severe axonal degeneration of the median nerve is an independent risk factor for prolonged IMV OR 4.9 (95%CI 1.1-21.5) p = 0.03, AUC of 0.774, (95%CI 0.66-0.88), p = < 0.001. CONCLUSIONS: Severe median nerve damage is an independent risk factor for prolonged mechanical ventilation.

Effect of the use of vitamin "E" in the diluent on the viability of ram sperm.

The sperm membrane is damaged in cryopreservation processes; this damage can be minimized using antioxidants such as vitamin E. The objective of the present study was to evaluate the effect of the addition of vitamin "E" on the viability of ram sperm during preservation processes. Two experiments were carried out; in the first, 32 ejaculates were used, which after evaluation were divided into two aliquots for processing; the first received Triladyl + vitamin "E" (T + E), and the second received only Triladyl (T); these aliquots were cooled and stored at 5 °C for 24 h. The viability (sperm motility, integrity, and membrane permeability) was evaluated at 0 and 24 h after dilution. In the second experiment, the same procedure was performed as experiment 1, except that the samples were also frozen, and the viability was evaluated at zero and 48 h post-freezing. Dependent variables were analyzed using mixed models in a split plot design. In experiment 1, the integrity and permeability of the sperm membrane was better in the group: "T + E" (P <0.05). In experiment 2, the vitamin significantly improved (P <0.05) the sperm viability. It is concluded that the addition of vitamin "E" improves sperm viability.

Ovarian dynamics of buffalo (Bubalus bubalis) synchronized with different hormonal protocols.

The study evaluates the effect of three hormonal protocols on ovarian dynamics and progesterone (P4) secretion of buffalo (Bubalus bubalis). Twenty-nine pluriparous Murrah buffaloes were used. The protocols were as follows: OVSYNCH (n = 10): 100 µg of gonadorelin (day 0), 500 µg of cloprostenol (day 7), and 100 µg of gonadorelin (day 9). CIDR+EB (intravaginal device (CIDR®) + estradiol benzoate; n = 10): CIDR plus 2 mg of EB (day 0), withdrew of CIDR, 500 µg of cloprostenol (day 7) and 1 mg of EB (day 8). CIDR+eCG (n = 9): CIDR plus 2 mg of EB (day 0), withdrew of CIDR, 500 µg of cloprostenol and 400 IU of eCG (day 7). Follicles were counted with an ultrasound and measured at 0, 24, and 54 h. The maximum follicle diameter and ovulation were evaluated at 70, 80, and 94 h after CIDR withdrew. Estrous was detected per 1 h three times daily. Blood samples were collected on days 0, 7, 10, 15, and 22 to determine P4 concentration. In CIDR+EB protocol, 50% of buffaloes presented estrous, at 69.6 h. All buffaloes ovulated. CIDR+eCG group had the shortest (69 h) ovulation time. No treatment differences for follicular population, maximum follicle diameter, and P4 concentration on days 7 and 10 (P > 0.05) were found. The P4 concentration in OVSYNCH and CIDR+eCG protocols were > 1 ng/ml, on days 15 and 22 (P < 0.05). There was no difference in ovarian activity; however, the P4 secretion was normal in the OVSYNCH and CIDR+eCG protocols compared to the CIDR+EB protocol.

Reproductive behavior in hair sheep rams under tropical conditions.

The reproductive efficiency of sheep herds depends to a great extent on the ram. Male reproductive evaluation allows to select for the best and eliminate those with reproductive problems. The objective was to evaluate the effect of breed and age group on the reproductive behavior of hair sheep rams in the tropics of Mexico. Pelibuey (n = 42), Blackbelly (n = 30), Dorper (n = 44), and Katahdin (n = 30) rams of two age groups: young (n = 74, 1-1.5 years old) and adult (n = 72, 2-4 years old) were evaluated. Serving capacity (10-min duration) and breeding soundness evaluation (BSE) tests were carried out in each ram. In the first test, rams were classified as suitable and unsuitable, and in the BSE test, they were classified as satisfactory, questionable, or unsatisfactory. The response variables were analyzed using chi-square test or analyses of variance (ANOVA). ANOVA included the fixed effects of breed, age group, and their interaction. In the serving capacity test, 79.5% of the rams were considered suitable, with no breed differences (P > 0.05). Adult rams (90.3%) had the highest proportion of suitable rams (P < 0.05). In the BSE test, 80.2% of the rams were satisfactory; only breed being significant (P < 0.05). Pelibuey breed had the highest proportion of satisfactory rams (91.4%). Breed × age interaction was no significant for any trait. After serving capacity and BSE tests, a high proportion of rams was found not suitable for reproduction (36.3%), which is expected to cause low fertility in the flock.

Type 2 diabetes mellitus BALB/c mice are more susceptible to granulomatous amoebic encephalitis: Immunohistochemical study.

Granulomatous amoebic encephalitis (GAE) is a chronic, difficult to resolve infection caused by amphizoic amoebae of the genus Acanthamoeba, which in most cases occurs in immunosuppressed persons or with chronic diseases such as diabetes. In this study, we describe the early events of A. culbertsoni infection of GAE in diabetic mice model. Diabetes was induced in male BALB/c mice, with a dose of streptozotocin (130 mg/kg). Healthy and diabetic mice were inoculated via intranasal with 1 × 106 trophozoites of A. culbertsoni. Then were sacrificed and fixed by perfusion at 24, 48, 72 and 96 h post-inoculation, the brains and nasopharyngeal meatus were processed to immunohistochemical analysis. Invasion of trophozoites in diabetic mice was significantly greater with respect to inoculated healthy mice. Trophozoites and scarce cysts were immunolocalized in respiratory epithelial adjacent bone tissue, olfactory nerve packets, Schwann cells and the epineurium base since early 24 h post-inoculation. After 48 h, trophozoites were observed in the respiratory epithelium, white matter of the brain, subcortical central cortex and nasopharyngeal associated lymphoid tissue (NALT). At 72 h, cysts and trophozoites were immunolocalized in the olfactory bulb with the presence of a low inflammatory infiltrate characterized by polymorphonuclear cells. Scarce amoebae were observed in the granular layer of the cerebellum without evidence of inflammation or tissue damage. No amoebas were observed at 96 h after inoculation, suggesting penetration to other tissues at this time. In line with this, no inflammatory infiltrate was observed in the surrounding tissues where the amoebae were immunolocalized, which could contribute to the rapid spread of infection, particularly in diabetic mice. All data suggest that trophozoites invade the tissues by separating the superficial cells, penetrating between the junctions without causing cytolytic effect in the adjacent cells and subsequently reaching the CNS, importantly, diabetes increases the susceptibility to amoebae infection, which could favor the GAE development.

In vivo CNS infection model of Acanthamoeba genotype T4: the early stages of infection lack presence of host inflammatory response and are a slow and contact-dependent process.

This study was developed in order to describe the early morphological events observed during the invasion of two pathogenic strains of Acanthamoeba (genotype T4); A. castellanii and A. culbertsoni, at the olfactory meatus and cerebral, pulmonary, renal, hepatic and splenic tissues levels, an in vivo invasion study. Histological and immunohistochemical description of the events at 24, 48, 72, and 96 h postintranasal inoculations of BALB/c mice was performed. A. castellanii showed a higher invasion rate than A. culbertsoni, which was only able to reach lung and brain tissue in the in vivo model. The current study supports previous evidence of lack of inflammatory response during the early stages of infection. Acanthamoeba invasion of the CNS and other organs is a slow and contact-dependent process. The early morphological events during the invasion of amoebae include the penetration of trophozoites into different epithelia: olfactory, respiratory, alveolar space, and renal tubule, which resemble the process of amoebae invasion described in corneal tissue. The data suggest that after reaching the nasal epithelium, trophozoites continued invasion, separating and lifting the most superficial cells, then migrating and penetrating between the cell junctions without causing a cytolytic effect on adjacent cells. These results reaffirm the idea that contact-dependent mechanisms are relevant for amoebae of Acanthamoeba genus regardless of the invasion site.

Comparison of effects of different statins on growth and steroidogenesis of rat ovarian theca-interstitial cells.

Statins are competitive inhibitors of 3-hydroxy-3-methyl-glutaryl-CoA reductase, the rate-limiting enzyme of the cellular production of cholesterol and other products of the mevalonate pathway. Statins exert hepatic and extrahepatic effects, modulating the function of various tissues and organs, including ovaries. Previously, we have demonstrated that simvastatin inhibited cellular proliferation and reduced androgen production by ovarian theca-interstitial cells. The above actions are of translational relevance to the most common endocrine disorder among women in reproductive age: polycystic ovary syndrome. However, different statins may have distinctly different profiles of effects on cholesterol and androgens. The present study was designed to compare the effects of several statins on growth and steroidogenesis of rat theca-interstitial cells. The cells were incubated in the absence (control) or in the presence of simvastatin, lovastatin, atorvastatin, or pravastatin. Assessment of effects of statins on cell growth was carried out by evaluation of DNA synthesis and by estimation of the number of viable cells. Effects on steroidogenesis were evaluated by quantification of steroid production and expression of mRNA for the key enzyme regulating androgen production: Cyp17a1. Among tested statins, simvastatin exerted the greatest inhibitory effects on all tested parameters. The rank order of the effects of the tested statins is as follows: simvastatin > lovastatin > atorvastatin ≥ pravastatin. While the lipophilicity is likely to play a major role in determining the ability of statins to act on nonhepatic cells, other factors unique to individual cell types are also likely to be relevant.

The combination of oral L-DOPA/rimonabant for effective dyskinesia treatment and cytological preservation in a rat model of Parkinson's disease and L-DOPA-induced dyskinesia.

Parkinson's disease is the second most prevalent neurodegenerative disease in the world. Its treatment is limited so far to the management of parkinsonian symptoms with L-DOPA (LD). The long-term use of LD is limited by the development of L-DOPA-induced dyskinesias and dystonia. However, recent studies have suggested that pharmacological targeting of the endocannabinoid system may potentially provide a valuable therapeutic tool to suppress these motor alterations. In the present study, we have explored the behavioral (L-DOPA-induced dyskinesias severity) and cytological (substantia nigra compacta neurons and striatum neuropil preservation) effects of the oral coadministration of LD and rimonabant, a selective antagonist of CB1 receptors, in the 6-hydroxydopamine rat model of Parkinson's disease. Oral coadministration of LD (30 mg/kg) and rimonabant (1 mg/kg) significantly decreased abnormal involuntary movements and dystonia, possibly through the conservation of some functional tyrosine hydroxylase-immunoreactive dopaminergic cells, which in turn translates into a well-preserved neuropil of a less denervated striatum. Our results provide anatomical evidence that long-term coadministration of LD with cannabinoid antagonist-based therapy may not only alleviate specific motor symptoms but also delay/arrest the degeneration of striatal and substantia nigra compacta cells.

Simvastatin reduces steroidogenesis by inhibiting Cyp17a1 gene expression in rat ovarian theca-interstitial cells.

Polycystic ovary syndrome (PCOS) is characterized by ovarian enlargement, theca-interstitial hyperplasia, and increased androgen production by theca cells. Previously, our group has demonstrated that statins (competitive inhibitors of 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway) reduce proliferation of theca-interstitial cells in vitro and decrease serum androgen levels in women with PCOS. The present study evaluated the effect of simvastatin on rat ovarian theca-interstitial cell steroidogenesis. Because actions of statins may be due to reduced cholesterol availability and/or isoprenylation of proteins, the present study also investigated whether steroidogenesis was affected by cell- and mitochondrion-permeable 22-hydroxycholesterol, isoprenylation substrates (farnesyl-pyrophosphate [FPP] and geranylgeranyl-pyrophosphate [GGPP]), as well as selective inhibitors of farnesyltransferase (FTI) and geranylgeranyltransferase (GGTI). Theca-interstitial cells were cultured for 12, 24, and 48 h with or without simvastatin, GGPP, FPP, FTI, GGTI, and/or 22-hydroxycholesterol. Simvastatin decreased androgen levels in a time- and concentration-dependent fashion. This inhibitory effect correlated with a decrease in mRNA levels of Cyp17a1, the gene encoding the key enzyme regulating androgen biosynthesis. After 48 h, GGPP alone and FPP alone had no effect on Cyp17a1 mRNA expression; however, the inhibitory action of simvastatin was partly abrogated by both GGPP and FPP. The present findings indicate that statin-induced reduction of androgen levels is likely due, at least in part, to the inhibition of isoprenylation, resulting in decreased expression of CYP17A1.

Alzheimer-like cell death after vanadium pentoxide inhalation.

Vanadium (V) toxicity depends on its oxidation state; it seems that vanadium pentoxide (V2O5) is the most toxic to the living cells. It has been reported that oral administration induces changes in motor activity and learning; in rats, I.P. administration increases lipid peroxidation levels in the cerebellum and the concentration of free radicals in the hippocampus and cerebellum. Mice that inhaled V2O5 presented a reduced number of tubulin+ in Leydig and Sertoli cells; it has also been reported that inhaled V2O5 induces loss of dendritic spines, necrosis, and hippocampus neuropil alterations; considering the direct consequence of the interaction of V with cytoskeletal components, makes us believe that V2O5 exposure could cause neuronal death in the hippocampus similar to that seen in Alzheimer disease. This work aimed to determine pyramidal hippocampal CA1 cytoskeletal alterations with Bielschowsky stain in rats exposed to V2O5. Male Wistar rats inhaled 0.02 M of V2O5 one h two times a week for two and six months. We found that rats, which inhaled V2O5 reached 56,57% of dead neurons after six months of inhalation; we recognize strong argyrophilic and collapsed somas and typical flame-shaped in all V-exposed rats hippocampus CA1 compared to controls. We also observe somatodendritic distortions. Axons and dendrites displayed thick dark bands replaced by noticeable thickening and nodosities and the cytoskeleton fibrillary proteins' linear traces. Our findings suggest that V2O5 inhalation induces Alzheimer-like cell death with evident cytoskeletal alterations.

Effects of resveratrol on proliferation and apoptosis in rat ovarian theca-interstitial cells.

Polycystic ovary syndrome (PCOS) is characterized by ovarian dysfunction and associated with ovarian theca-interstitial (T-I) cell hyperplasia, hyperinsulinemia, systemic inflammation and oxidative stress. This in vitro study tested whether rat T-I cell growth with or without insulin can be altered by resveratrol, a natural polyphenol with anti-carcinogenic, anti-inflammatory, anti-proliferative and antioxidant properties. Rat T-I cells were cultured with and without resveratrol and/or insulin, and the effects on DNA synthesis, number of viable cells and markers of apoptosis were evaluated. Resveratrol alone induced a potent concentration-dependent inhibition of cell growth by inhibiting DNA synthesis, decreasing the number of viable cells and increasing the activity of executioner caspases 3 and 7; these effects of resveratrol counteracted the pro-proliferative and anti-apoptotic effects of insulin. Immunofluorescence analysis of cells incubated with resveratrol showed concentration- and time-dependent morphological changes consistent with apoptosis. The present findings indicate that resveratrol promotes apoptosis to reduce rat T-I cell growth in vitro as well as inhibiting insulin-induced rat T-I cell growth. This suggests a possibility that resveratrol and/or mechanisms mediating its effect may be relevant to the development of novel treatments for PCOS, which is characterized by both excessive ovarian mesenchyma growth and hyperinsulinemia.

Protracted hyperthermia and delayed rhabdomyolysis in ecstasy toxicity: A case report.

RATIONALE: Despite toxicity and unpredictable adverse effects, ecstasy use has increased in the United States. Onset of hyperpyrexia, rhabdomyolysis, disseminated intravascular coagulation (DIC), among other symptoms, occurs within hours of ingestion. Moreover, patients who experience hyperpyrexia, altered mental status, DIC, and multiorgan failure, rarely survive. This case presents a chronic ecstasy user whose symptoms would have predicted mortality. The report demonstrates a patient who experiences protracted hyperthermia, with delayed rhabdomyolysis and DIC. In addition, his peak creatine kinase (CK) of 409,440 U/L was far greater than the expected 30,000 to 100,000 U/L, being the second largest CK recorded in a survivor. PATIENT CONCERNS: This case report presents a 20-year-old man who presented to the emergency department after experiencing a severe reaction to ecstasy. He was a chronic user who took his baseline dosage while performing at a music event. He experienced hyperpyrexia immediately (106.5°F) while becoming stiff and unresponsive. Before emergency medical service arrival, his friends placed cold compresses on the patient and rested him in an ice filled bathtub. DIAGNOSES: Per history from patient's friends and toxicology results, the patient was diagnosed with ecstasy overdose, which evolved to include protracted hyperthermia and delayed rhabdomyolysis. INTERVENTIONS: Due to a Glasgow coma scale score of 5, he was intubated and sedated with a propofol maintenance. Hyperpyrexia resolved (temperature dropped to 99.1°F) after start of propofol maintenance. He was extubated after 24 hours, upon which he experienced hyperthermia (101.4°F at 48 hours), delayed rhabdomyolysis, and DIC (onset at 37 hours). He remained in hyperthermia for 120 hours until carvedilol permanently returned his temperature to baseline. His plasma CK reached a peak of 409,440 U/L at 35 hours. OUTCOMES: After primary management with intravenous fluids, the patient returned to baseline health without any consequences and was discharged after 8 days. A follow-up of 3 months postdischarge revealed no complications or disability. LESSONS: Clinically, the case highlights how physicians should be aware of the unusual time course adverse effects of ecstasy can have. Lastly, as intensity and duration of hyperpyrexia are predictors of mortality, our case indicates maintenance of sedation with propofol and use of oral carvedilol; both are efficacious for temperature reduction in ecstasy toxicity.

Author Correction: Time-restricted feeding restores muscle function in Drosophila models of obesity and circadian-rhythm disruption.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Role of isoprenylation in simvastatin-induced inhibition of ovarian theca-interstitial growth in the rat.

Statins are competitive inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway. The pleiotropic effects of statins may be due to inhibition of cholesterol synthesis, as well as decreased availability of several biologically important intermediate components of the mevalonate pathway, including two substrates for isoprenylation (farnesyl pyrophosphate [FPP] and geranylgeranyl pyrophosphate [GGPP]). Recently, we demonstrated statin-induced inhibition of ovarian theca-interstitial cell proliferation in vitro, as well as reduction of testosterone levels in women with polycystic ovary syndrome (PCOS). This study evaluates the relative contribution of inhibition of isoprenylation and/or cholesterol availability to the modulation of theca-interstitial proliferation. Rat theca-interstitial cells were cultured in chemically defined media with or without simvastatin, FPP, GGPP, squalene, and/or two membrane-permeable forms of cholesterol (25-hydroxycholesterol and 22-hydroxycholesterol). Simvastatin inhibited DNA synthesis and the count of viable cells. The effects of simvastatin were partly abrogated by FPP and GGPP but not by squalene or cholesterol. Inhibition of farnesyl transferase and geranylgeranyl transferase reduced cell proliferation. The present findings indicate that simvastatin inhibits proliferation of theca-interstitial cells, at least in part, by reduction of isoprenylation. These observations provide likely mechanisms explaining clinically observed improvement of ovarian hyperandrogenism in women with PCOS.

Time-restricted feeding restores muscle function in Drosophila models of obesity and circadian-rhythm disruption.

Pathological obesity can result from genetic predisposition, obesogenic diet, and circadian rhythm disruption. Obesity compromises function of muscle, which accounts for a majority of body mass. Behavioral intervention that can counteract obesity arising from genetic, diet or circadian disruption and can improve muscle function holds untapped potential to combat the obesity epidemic. Here we show that Drosophila melanogaster (fruit fly) subject to obesogenic challenges exhibits metabolic disease phenotypes in skeletal muscle; sarcomere disorganization, mitochondrial deformation, upregulation of Phospho-AKT level, aberrant intramuscular lipid infiltration, and insulin resistance. Imposing time-restricted feeding (TRF) paradigm in which flies were fed for 12 h during the day counteracts obesity-induced dysmetabolism and improves muscle performance by suppressing intramuscular fat deposits, Phospho-AKT level, mitochondrial aberrations, and markers of insulin resistance. Importantly, TRF was effective even in an irregular lighting schedule mimicking shiftwork. Hence, TRF is an effective dietary intervention for combating metabolic dysfunction arising from multiple causes.

Time course changes of the striatum neuropil after unilateral dopamine depletion and the usefulness of the contralateral striatum as a control structure.

INTRODUCTION: After unilateral dopamine depletion, some ipsilateral alterations occur and the contralateral structure has been utilized as control. OBJECTIVE: Our aim is to analyse the evolution of the ultrastructural alterations of the ipsilateral and contralateral striata of the 6-hydroxydopamine lesioned rats to demonstrate that the contralateral striatum should not be used as control structure. METHODS: After the surgery and the rotation behavior evaluation, animals were killed from 3 to 120 days after lesioning, and their striata were compared with those of aged rats. RESULTS: The ultrastructural analysis shows increased diameter of the synaptic ending in ipsilateral (since the third day) and contralateral striata (since day 30) and an increase in perforated synaptic contacts. CONCLUSION: Our data suggest that the contralateral striatum should not be taken as control structure at least after 20-30 days after lesioning, as the alterations found here may result in wrong interpretations when comparing with the ipsilateral-lesioned one.

Effect of season on follicular population, quality and nuclear maturation of bovine oocytes under tropical conditions.

The aim was to determine the effect of season of the year and the presence of a corpus luteum (CL) on follicular population (FP) and the quality of the oocytes, and of season on nuclear maturation of the bovine oocytes under tropical conditions. Three seasons were evaluated: hot-dry (March-June), hot-humid (July-October) and fresh-humid (November-February). In a first study, 1112 bovine ovaries were obtained from a local slaughterhouse. Follicles were classified as small (≤4mm), middle (4.1-8mm) and large (≥8.1mm); and the maximum diameter of the follicle (MDF) and CL (MDCL) were also recorded. The oocytes were collected by aspiration and classified as viable (grade I and II) and damaged (grade III and IV). In the second study, 2261 viable oocytes were matured in vitro, and then fixed and stained with Lacmoid to classify the stage of development as mature (metaphase II), immature or degenerate. Data were analyzed using analysis of variance and chi-square procedures. The largest FP of large follicles (0.67), MDF (1.18mm), MDCL (1.87mm), and the highest proportion of viable oocytes (34.19%) were obtained during the hot-humid season (P<0.05). The ovaries without CL had the greatest FP (10.34) with more viable oocytes (24.44%). The highest proportion of mature oocytes (76.92%) was also obtained in the hot-humid season. In conclusion, season influenced FP, MDF, MDCL, and the quality and nuclear maturation of oocytes. The presence of a CL in the ovary resulted in a decrease of FP and viability of oocytes.

Interactions of ethanol and folate deficiency in development of alcoholic liver disease in the micropig.

Folate deficiency is present in most patients with alcoholic liver disease (ALD), whereas folate regulates and alcoholism perturbs intrahepatic methionine metabolism, and S-adenosyl-methionine prevents the development of experimental ALD. Our studies explored the hypothesis that abnormal methionine metabolism is exacerbated by folate deficiency and promotes the development of ALD in the setting of chronic ethanol exposure. Using the micropig animal model, dietary combinations of folate deficiency and a diet containing 40% of kcal as ethanol were followed by measurements of hepatic methionine metabolism and indices of ALD. Alcoholic liver injury, expressed as steatohepatitis in terminal 14 week liver specimens, was evident in micropigs fed the combined ethanol containing and folate deficient diet but not in micropigs fed each diet separately. Perturbations of methionine metabolism included decreased hepatic S-adenosylmethionine and glutathione with increased products of DNA and lipid oxidation. Thus, the development of ALD is linked to abnormal methionine metabolism and is accelerated in the presence of folate deficiency.

Folate deficiency, methionine metabolism, and alcoholic liver disease.

Methionine metabolism is regulated by folate, and both folate deficiency and abnormal hepatic methionine metabolism are recognized features of alcoholic liver disease (ALD). Previously, histological features of ALD were induced in castrated male micropigs fed diets containing ethanol at 40% of kilocalories for 12 months, whereas in male micropigs fed the same diets for 12 months abnormal methionine metabolism and hepatocellular apoptosis developed. Folate deficiency may promote the development of ALD by accentuating abnormal methionine metabolism. Intact male micropigs received eucaloric diets that were folate sufficient, folate deficient, or each containing 40% of kilocalories as ethanol for 14 weeks. Folate deficiency alone reduced hepatic folates by one half, and ethanol feeding alone reduced methionine synthase, S-adenosylmethionine (SAM), and glutathione (GSH) levels and elevated plasma malondialdehyde (MDA) levels. The combined regimen elevated plasma homocysteine, hepatic S-adenosylhomocysteine (SAH), urinary 8-hydroxy-2-deoxyguanosine (oxy(8)dG), an index of DNA oxidation, and serum aspartate aminotransferase (AST) levels. Terminal hepatic histopathologic characteristics included typical features of steatonecrosis and focal inflammation in pigs fed the combined diet, with no changes in the other groups. Hepatic SAM levels correlated with those of GSH, whereas urinary oxy(8)dG and plasma MDA levels correlated with the SAM:SAH ratio and to hepatic GSH. The results demonstrate the linkage of abnormal methionine metabolism to products of DNA and lipid oxidation and to liver injury. The finding of steatonecrosis and focal inflammation only in the combined diet group supports the suggestion that folate deficiency promotes and folate sufficiency protects against the early onset of methionine cycle-mediated ALD.

Resveratrol potentiates effects of simvastatin on inhibition of rat ovarian theca-interstitial cells steroidogenesis.

BACKGROUND: Polycystic ovary syndrome (PCOS) is characterized by ovarian enlargement, hyperplastic theca compartment and increased androgen production due to, at least in part, excessive expression of several key genes involved in steroidogenesis. Previously, our group has demonstrated that simvastatin, competitive inhibitor of 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA reductase), a rate-limiting step of the mevalonate pathway, reduces rat-theca interstitial cell steroidogenesis by inhibiting Cyp17a1 gene expression, the key enzyme of the androgen biosynthesis pathway. Recently, we demonstrated that resveratrol, a bioflavonoid abundant in red grapes, decreases rat theca-interstitial cell steroidogenesis and this suppressive effect is mediated through mechanisms independent of the mevalonate pathway. The present study evaluated the effect of combining simvastatin and resveratrol treatments on rat theca-interstitial cell steroidogenesis. METHODS: Rat theca-interstitial cells isolated from 30 day-old female rats were cultured for up to 48 h with or without simvastatin (1 µM) and/or resveratrol (3-10 µM). Steroidogenic enzymes gene expression was evaluated by quantitative real time PCR and steroid levels were measured by liquid chromatography-mass spectrometry. Comparisons between groups were performed using ANOVA and Tukey test. RESULTS: Resveratrol potentiated inhibitory effects of simvastatin on androstenedione and androsterone production in theca-interstitial cells. This suppressive effect correlated with profound inhibition in Cyp17a1 mRNA expression in the presence of a combination of resveratrol and simvastatin. CONCLUSIONS: The present findings indicate that resveratrol potentiates the simvastatin-induced inhibitory effect on theca-interstitial cell androgen production, raising the possibility of development of novel treatments of PCOS.