Follicular development, morphological integrity, and oxidative stress in bovine preantral follicles cultured in vitro with ascorbic acid.

The aim of this study was to evaluate the follicular development, morphological integrity, and oxidative stress of preantral ovarian follicles from Bos taurus indicus females grown in vitro with ascorbic acid. Ovaries (n = 20) from Bos taurus indicus females were collected, fragmented, and were cultured in vitro for 6 or 12 days in minimum essential medium (MEM), or MEM supplemented with 50 or 100 ng/ml ascorbic acid, with an extracellular matrix of agarose gel, in an incubator at 38.5°C; every 2 days, 100% of the culture medium was replaced. The data were analyzed using the chi-squared test and/or Fisher's exact test. In the event of a significant effect, the proportions were compared using a 2 × 2 proportion test. The oxidative stress analysis data were submitted to analysis of variance followed by the Bonferroni test. Values were considered significant when P ≤ 0.05. The addition of 100 ng/ml of ascorbic acid to the in vitro culture medium of preantral ovarian follicles from bovine females promoted follicular development, was efficient in maintaining morphological integrity, as well as the stability of reactive oxygen species, after 6 days of in vitro culture.

Culture of preantral ovarian follicles of Bos taurus indicus with alpha-lipoic acid.

The aim of this study was to evaluate follicular development, morphological integrity, and antioxidant potential of preantral ovarian follicles from Bos taurus indicus females grown in vitro with alpha-lipoic acid. Ovaries (n = 24) of Bos taurus indicus (n = 12) females were collected during slaughter and fragmented. A randomly obtained fragment from each pair of ovaries was fixed in Bouin (non-cultivated control; D0). These fragments were intended for classical histology (morphology and evaluation of follicular growth), and a fragment from each pair of ovaries was frozen at -80°C (non-cultivated control; D0), and assigned for analysis of oxidative stress [thiobarbituric acid reactive substances (TBARS), nitroblue tetrazolium (NBT), and ferric reducing antioxidant power (FRAP)]. The remaining fragments were cultured in vitro for 6 (D6) or 12 (D12) days, containing only minimum essential medium (MEM) or MEM supplemented with alpha-lipoic acid (50, 100, or 250 ng/ml), on an extracellular matrix of agarose gel, in an oven at 38.5ºC. Every 2 days, 100% of the culture medium was replaced. Supplementation with 100 ng/ml was effective for maintaining follicular integrity after 6 days of culture (primordial: 51.28%; development: 36.88%; P < 0.0001). There was no difference (P > 0.05) between treatments compared with the non-cultivated control treatment (D0), using the NBT and TBARS assays. Therefore, supplementation of the in vitro culture medium of bovine preantral ovarian follicles with a concentration of 100 ng/ml of alpha-lipoic acid at 6 days of culture was effective for maintaining follicular integrity and, after 6 days, maintaining stable levels of reactive oxygen species.

Secreted Osteoclastogenic Factor of Activated T Cells (SOFAT) Is Associated With Rheumatoid Arthritis and Joint Pain: Initial Evidences of a New Pathway.

Rheumatoid arthritis (RA) has an inflammatory milieu in the synovial compartment, which is regulated by a complex cytokine and chemokine network that induces continuously degenerative and inflammatory reactions. The secreted osteoclastogenic factor of activated T cells (SOFAT) is a unique cytokine and represents an alternative pathway for osteoclast activation. In this study, we examined whether SOFAT is able to induce joint pain and investigated the presence of SOFAT in a Collagen-induced Arthritis (CIA) model and in human subjects. Here, we found that an intra-articular stimulation with SOFAT (1, 10, 100, or 1,000 ng/10 µl) in the knee joint significantly decreases the mechanical threshold in the hind paw of mice (p < 0.05). Moreover, after a second injection of SOFAT, the mechanical threshold decrease was sustained for up to 8 days (p < 0.05). In the CIA model, the immunohistochemical assay of knee joint showed positivity stained for SOFAT, and the mRNA and protein expression of SOFAT were significantly higher in the affected-group (p < 0.05). Besides, the mRNA of RANKL, IL-1ß, IL-6, and IL-15 were significantly higher in the affected-group (p < 0.05). Finally, SOFAT was detected in the synovial fluid of RA patients, but not in OA patients (p < 0.05). In conclusion, SOFAT is up regulated in inflammatory milieu such as RA but not in non-inflammatory OA. SOFAT may be a novel molecule in the complex inflammatory phenotype of RA.

Safety and efficacy of superior turbinate biopsies as a source of olfactory epithelium appropriate for morphological analysis.

PURPOSE: There is no standardized approach for preserving olfactory function in the side of the nose where biopsy of the olfactory epithelium (OE) is performed. Moreover, a gold standard technique for obtaining human OE in vivo is still lacking. We determined the efficacy of obtaining good-quality OE specimens suitable for pathological analysis from the lower half of the superior turbinate and verified the safety of this procedure in maintaining bilateral and unilateral olfactory function. METHODS: In 21 individuals without olfactory complaints and who had undergone septoplasty and inferior turbinectomy OE biopsy was made during septoplasty. Olfactory function, both unilateral and bilateral, was assessed using the University of Pennsylvania Smell Identification Test (UPSIT) before and 1 month after the procedure. Specimens were marked with the olfactory marker protein for confirmation of OE presence. RESULTS: Ninety percent of the samples contained OE, although clear histological characterization was possible from only 62%. There was no deterioration of UPSIT scores either bilaterally or unilaterally on the side of the biopsy. Patients also maintained the ability to identify individual odorants. CONCLUSION: Biopsies of the lower half of the superior turbinate do not affect olfactory function and show strong efficacy in yielding OE tissue and moderate efficacy for yielding tissue appropriate for morphological analysis. Future studies are needed to assess the safety of this procedure in other OE regions.

Depressed female smokers have higher levels of soluble tumor necrosis factor receptor 1.

AIM: To examine clinical and biomarkers in depressed female smokers, in order to better clarify the process that link mood disorders, childhood trauma and smoking in women. METHODS: The clinical sample comprised women with unipolar or bipolar depression, divided into subgroups of smokers and never-smoker. The control groups comprised two subgroups non-depressed women, separated into smokers and never-smokers. A structured questionnaire was used to assess socio-demographic and clinical data. The following scales were used: 17-item version Hamilton Depression Rating Scale, Hamilton Anxiety Rating scale (HAM-A), Sheehan disability scale, the Child Trauma Questionnaire. The following biomarkers were investigated: lipid profile, including total cholesterol, high-density lipoprotein cholesterol (HDLc), low-density lipoprotein cholesterol, triglycerides the Castelli's Risk indexes I and II; and cytokines, including interleukins (IL)-1ß, IL-6, IL-10, IL-12, soluble tumor necrosis factor receptor 1 (sTNF-R1). RESULTS: Depressed female smokers showed a number of significant positive correlations: emotional neglect and sTNF-R1 (p = 0.02); waist circumference and sTNF-R1 (p = 0.001); body mass index and sTNF-R1 (p < 0.01); HAM-A and sTNF-R1 (p = 0.03); IL-1ß and sTNF-R1 (p < 0.01); IL-10 and sTNF-R1 (p = 0.001); IL-12 and sTNF-R1 (p < 0.01);Castelli index I and sTNF-R1 (p < 0.01); Castelli index II and sTNF-R1 (p < 0.01); and a significantly negative correlation between HDLc and sTNF-R1(p = 0.014). CONCLUSION: This study suggests that depressed female smokers who experienced more childhood trauma and had more anxiety symptoms are associated with the activation of inflammatory processes and alterations in components of lipid profile.

The effect of intrathecal gabapentin on neuropathic pain is independent of the integrity of the dorsolateral funiculus in rats.

AIM: This study evaluates the contribution of inhibitory pain pathways that descend to the spinal cord through the dorsolateral funiculus (DLF) on the effect of intrathecal gabapentin against spinal nerve ligation (SNL)-induced behavioral hypersensitivity to mechanical stimulation in rats. MAIN METHOD: Rats were submitted to a sham or complete ligation of the right L5 and L6 spinal nerves and a sham or complete DLF lesion. Next, the changes induced by intrathecal administration of gabapentin on the paw withdrawal threshold of rats to mechanical stimulation were evaluated electronically. KEY FINDINGS: Intrathecal gabapentin (200µg/5µl) that was injected 2 or 7days after surgery fully inhibited the SNL-induced behavioral hypersensitivity to mechanical stimulation in sham DLF-lesioned rats; gabapentin was effective against the SNL-induced behavioral hypersensitivity to mechanical stimulation also in DLF-lesioned rats. SIGNIFICANCE: The effect of intrathecally administered gabapentin against SNL-induced behavioral hypersensitivity to mechanical stimulation in rats does not depend on the activation of nerve fibers that descend to the spinal cord via the DLF.

Stimulation of the occipital or retrosplenial cortex reduces incision pain in rats.

The electrical stimulation of the occipital (OC) or retrosplenial (RSC) cortex produces antinociception in the rat tail-flick and formalin tests. This study examined the antinociceptive effects of stimulating the OC or RSC in a rat model of post-incision pain. The involvement of the anterior pretectal nucleus (APtN) as intermediary for the effect of OC or RSC stimulation was also evaluated because the OC and RSC send inputs to the APtN, which is implicated in antinociception and nociception. It is shown that a 15-s period of electrical stimulation of the OC or RSC significantly reduced post-incision pain for less than 10 min and at least 15 min, respectively. The injection of 2% lidocaine (0.25 µl), naloxone (10 ng/0.25 µl), methysergide (40 pg/0.25 µl), or atropine (100 ng/0.25 µl) into the APtN produced a further increase in post-incision pain. The effect of RSC stimulation was shorter and less intense in rats pretreated with lidocaine, methysergide or naloxone. The effect of OC stimulation was shorter and less intense in lidocaine-treated rats, but remained unchanged in rats pretreated with methysergide or naloxone in the APtN. The effects of stimulating the OC or RSC were not changed in rats treated with atropine. We conclude that stimulation-induced antinociception from the RSC or OC in rat post-incision pain activates distinct descending pain inhibitory pathways. The pathway activated from the RSC utilizes serotonergic and opioid mediation in the APtN, whereas stimulation of the OC utilizes a non-serotonergic, non-cholinergic and non-opioid mediation in the same nucleus.

Stimulation-produced analgesia from the occipital or retrosplenial cortex of rats involves serotonergic and opioid mechanisms in the anterior pretectal nucleus.

UNLABELLED: The electrical stimulation of the occipital (OC) or retrosplenial (RSC) cortex produces antinociception in the rat tail-flick test. These cortices send inputs to the anterior pretectal nucleus (APtN) which is implicated in antinociception and nociception. At least muscarinic cholinergic, opioid, and serotonergic mechanisms in the APtN are involved in stimulation-produced antinociception (SPA) from the nucleus. In this study, the injection of 2% lidocaine (.25 µL) or methysergide (40 and 80 ng/.25 µL) into the APtN reduced the duration but did not change the intensity of SPA from the OC, whereas both duration and intensity of SPA from the RSC were significantly reduced in rats treated with lidocaine or naloxone (10 and 50 ng/.25 µL), injected into the APtN. Naloxone or methysegide injected into the APtN was ineffective against SPA from the OC or RSC, respectively. Atropine (100 ng/.25 µL) injected into the APtN was ineffective against SPA from either the OC or RSC. We conclude that the APtN acts as an intermediary for separate descending pain inhibitory pathways activated from the OC and RSC, utilizing at least serotonin and endogenous opioid as mediators in the nucleus. PERSPECTIVE: Stimulation-induced antinociception from the retrosplenial or occipital cortex in the rat tail-flick test depends on the activation of separate descending pain inhibitory pathways that utilize the APtN as a relay station.