Effect of Melatonin on the Content of CD3low and CD3hi T Cells in the Thymus of Mice with Functional Pinealectomy.

Continuous lighting for 14 days (functional pinealectomy model) leads to a decrease in the relative number of CD3low and CD3hi T lymphocytes and the CD3low/CD3hi ratio in the thymus of C57BL/6 mice. Intragastric administration of melatonin in physiological doses (1 mg/kg body weight, 14 days) against the background of functional pinealectomy restores the percentage of CD3low and CD3hi thymocytes and CD3low/CD3hi ratio to the control values. Hence, prolonged continuous illumination inhibits the differentiation and maturation of young thymocytes into mature forms, while melatonin treatment helps to compensate the effects of functional pinealectomy triggering cell proliferation in the thymus from the earliest stages of proliferation and differentiation of T cells. Thus, melatonin has immunotropic properties and can be used for correction of the consequences of functional pinealectomy.

Neck Pain and Headache After Pinealectomy: Improvement with Multimodal Chiropractic Therapies.

BACKGROUND Pineal gland tumors are rare central nervous system tumors, and while neck pain and headaches may be common among those who have had these tumors removed, there is little research regarding management of these symptoms. CASE REPORT A 45-year-old man with a history of pineal germinoma treated with pinealectomy, chemotherapy, radiation therapy, and ventriculoperitoneal shunt placement at age 21 presented with chronic neck pain and headaches, which initially improved following his surgery and concurrent therapies, yet progressively worsened over the following years. He required thyroid and testosterone medication because of radiation-induced hypopituitarism, yet was employed, and until recently, active with playing tennis. He had previously seen his primary care provider, orthopedist, and neurologist, and had been cleared of severe pathology via brain magnetic resonance imaging and was referred to the chiropractor. On examination, the patient had severely limited passive cervical spine range of motion, yet hat no neurologic deficits, and radiographs showed mild cervical spondylosis and cervicothoracic scoliosis. His history and presentation were suggestive of radiation-induced fibrosis. The patient's neck pain, headaches, and quality of life improved with multimodal treatments including spinal and soft-tissue manipulation, stretches, and yoga. CONCLUSIONS This case illustrates long-term sequelae of a pineal gland tumor and its treatment, including neck pain and headache, and improvement with multimodal chiropractic therapies. Despite the success in this case, these results are not broadly generalizable. Further research is needed to understand the natural history of symptoms and effectiveness of multimodal therapies among patients who have had pineal tumor surgery.

Pinealectomy in Rats.

The pinealectomy technique consists of the surgical removal of the superficial pineal gland. This procedure allows the ablation of circulating indoles produced by this gland. Withdrawal of systemic melatonin, a pineal hormone, affects animal circadian rhythms and induces several physiological changes that are the subject of many investigations. In this chapter, we describe the pinealectomy protocol adapted to rats. We describe the animal placement on the stereotaxic fixation system, and the procedure for the pineal gland removal and animal recovery from surgery.

Light-Induced Functional Pinealectomy: Expression of MT2 Receptors in Liver Cells of C57BL/6 Mice after Melatonin Treatment.

We performed an immunohistochemical study of MT2 melatonin receptor expression in the liver of C57BL/6 mice with modeled light-induced functional pinealectomy and after melatonin administration by the indirect avidin-biotin peroxidase ABC method. The animals were kept for 14 days under constant lighting. Intragastric administration of melatonin in physiological doses (1 mg/kg body weight for 14 days) to mice with light-induced functional pinealectomy resulted in a 2-fold increase in the relative expression area of MT2 receptors in liver cells in comparison with that in animals kept under standard lighting conditions, 24-h lighting for 14 days, or 24-h lighting receiving placebo (intragastric administration of 200 ml distilled water). Melatonin treatment had practically no effect on MT2 staining intensity. Our results attest to the important role of MT2 receptors in melatonin synthesis disorders and can serve as the basis for the development of therapeutic strategies aimed at melatonin receptors.

Effect of Melatonin on the Expression of LYVE-1 Receptor in Liver Sinusoidal Endothelial Cells of Mice with the Light-Induced Functional Pinealectomy Model.

Intragastric administration of melatonin in physiological doses (1 mg/kg body weight) for 14 days to C57BL/6 mice with light-induced functional pinealectomy model (24-h lighting for 14 days) results in an increase in the LYVE-1 expression area by 2.4 times and a significant increase in receptor concentration (1.6% decrease in staining brightness) in liver sinusoidal endothelial cells in comparison with animals kept under continuous lighting and not treated with the hormone, which indicates the formation of stability of the endothelial barrier in the organ. Melatonin treatment also enhanced lymphatic drainage in all it links (including interstitial non-vascular pathways and lymphatic vessels) and improved structural and functional parameters of blood circulation and lymph flow in the organ, which created conditions for reducing metabolic load on structural elements of the liver.

Constant light exposure and/or pinealectomy increases susceptibility to trichloroethylene-induced hepatotoxicity and liver cancer in male mice.

Exposure to light at night, pineal gland impairment, and the environmental pollutant trichloroethylene (TCE) have serious implications for health and contribute to illness, including liver cancer. The adverse effect of the association of continuous exposure to light with decreased melatonin levels and TCE-induced toxicity is not disclosed in target organs. This work explored the role of light and pineal impairment in increasing susceptibility to liver toxicity and cancer upon exposure to TCE. Male albino mice were divided into groups as follows: control group (12-h light/12-h dark cycle), constant light (24-h light), pinealectomized (Pnx) mice, sham surgically treated group, TCE-treated groups subjected to two doses (500 and 1000 mg/kg) at two different light regimens, and combination of Pnx and TCE-treated mice kept at a 12-h light/12-h dark cycle. Melatonin levels were significantly decreased in both Pnx mice and TCE-treated animals at both light regimens. Aspartate transaminase, alanine aminotransferase, activities, and serum bilirubin levels were significantly elevated, whereas albumin levels were markedly decreased in Pnx mice, TCE-treated mice, and the combination group. Histopathological investigations reflected changes in liver function parameters indicating liver injury and induction of cancer. These effects were accompanied by significant increase of the liver cancer biomarker alpha-fetoprotein and the expression of the metastatic markers CD44, TGFß-1, and VEGF, along with increased oxidative stress indicators and inflammatory cytokines (IL-6, IL-1ß, and TNF-α) in both Pnx and TCE-treated mice and the combination group at both light regimens. Taken together, our findings indicated that low melatonin levels, exposure to constant light, and the combination of both factors increases susceptibility to the toxic and carcinogenic effects of TCE on the liver.

Influence of Pinealectomy and Long-term Melatonin Administration on Inflammation and Oxidative Stress in Experimental Gouty Arthritis.

Gout is an inflammatory arthritis characterized by the deposition of monosodium urate (MSU) crystals in the joints or soft tissue. MSU crystals are potent inflammation inducers. Melatonin (MLT) is a powerful endogenous anti-inflammatory agent and effective in reducing cellular damage. In the present study, possible underlying mechanisms associated with anti-inflammatory and antioxidative effects were investigated in rats with gouty arthritis and melatonin deprivation treated with MLT. Fifty-six rats were divided into seven groups: control, sham control, pinealectomy (PNX), MSU (on the 30th day, single-dose 20 mg/ml, intraperitoneal), MSU + MLT (10 mg/kg/day for 30 days, intraperitoneal), MSU + PINX and MSU + PINX + MLT. PNX procedure was performed on the first day of the study. As compared to the controls, the results showed that MSU administration caused significant increases in oxidative stress parameters (malondialdehyde and total oxidant status). Besides, significant decreases in antioxidant defense systems (glutathione, superoxide dismutase and total antioxidant status) were observed. A statistically significant increase was found in the mean histopathological damage score in the groups that received MSU injection. It was found that histopathological changes were significantly reduced in the MSU + MLT group given MLT. In our study, it was determined that many histopathological changes, as well as swelling and temperature increase in the joint, which are markers of inflammation, were significantly reduced with MLT supplementation. These results suggest that melatonin ameliorates MSU-induced gout in the rat through inhibition of oxidative stress and proinflammatory cytokine production.

Treatment with crocin suppresses diabetic nephropathy progression via modulating TGF-ß1 and oxidative stress in an experimental model of pinealectomized diabetic rats.

One of the most common complications of diabetes is diabetic nephropathy (DN). Uncontrolled hyperglycemia leads to histopathologic alterations in the kidney that prevent normal renal function. This study aimed to explore the effects of crocin treatment via virtue of its numerous beneficial properties in streptozotocin-induced pinealectomized diabetic rats. The pinealectomy procedure was conducted on the first day of the study. On the 30th day following pinealectomy, streptozotocin (STZ) (50 mg/kg) was administered intraperitoneally in Wistar rats for induction of diabetes. Diabetes was confirmed on the 3rd day following STZ administration by determining the glucose levels. Daily crocin treatment intraperitoneally for 15 days (50 mg/kg) ameliorated impaired renal oxidant/antioxidant balance, reduced TGF-ß1 immuno-staining around tubules, and promoted improvement of renal architecture. Moreover, crocin administration improved altered renal function parameters, including serum Cr and BUN, and also increased creatinine clearance. In conclusion, the protective effects of crocin on diabetic nephropathy might be associated with its powerful antioxidant properties, its ability to improve tissue antioxidant status, and its ability to prevent inflammatory pathways.

The influence of functional pinealectomy and exogenous melatonin application on healing of a burr hole in adult rat calvaria: a histological and immunohistochemical study.

BACKGROUND: Even today, repair of the cranial defects still represents a significant challenge in neurosurgery and various options have been used for their reconstruction to date. However, there are very few studies investigating the effects of exogenous administration of melatonin (MEL) as an agent that promotes bone regeneration. The goal of this study was to investigate the effects of functional pinealectomy (Px) and exogenous MEL administration on the bone repair properties and surrounding connective tissue alterations in a rat calvaria model. MATERIALS AND METHODS: The total of 30 adult female Wistar-Albino rats was randomly divided into three groups (n = 10): control group (CO; 12 h light/12 h dark exposure), functional Px group (24 h light exposure, light-induced functional Px), and Px+MEL group (light-induced Px + MEL, 20 mg/kg/day for 12 weeks). Critical-sized burr-hole defects (diameter: 3.0 mm) were surgically created by a single operator in the calvarium of all rats, using an electric drill. Animals in Px+MEL group received MEL 20 mg/kg/day for 12 weeks. At the end of the study, bone healing and connective tissue alterations surrounding drilled defect area in the rat calvaria were determined in haematoxylin-eosin-stained and Mallory Azan slices applied in anti-bone sialoprotein. Image Pro Express 4.5 programme was used for histomorphometric calculation of areas of new bone and fibrotic tissue. Normality control was performed by Shapiro-Wilk test. Variance homogeneities were examined by Shapiro-Wilk and Levene tests; Tukey HSD test was used as a post hoc method since there was no homogeneity problem. All hypothesis tests were performed at the 0.05 significance level. RESULTS: Histological analysis showed that the bone repair process in the Px+MEL group was similar to that of the CO group, whereas the functional Px group showed a delay. Histomorphometrically, it was found that the Px group had the largest hole diameter and the most fibrotic scar area, although no binary statistical significance was found between the CO and Px+MEL groups (p = 0.910). In terms of vascularisation, it was observed that the most vascular structure was found in the Px+MEL group among the scar tissue and ossification areas, while the vascularisation was the least in the Px group (p < 0.001). CONCLUSIONS: Our findings revealed that bone repair process was impaired in functional Px group, but exogenous MEL replacement was able to restore this response. Thus, it is concluded that utilisation of MEL may improve the bone repair in calvarial defects.

Effects of pinealectomy and crocin treatment on rats with isoproterenol-induced myocardial infarction.

The present study aimed to analyze the effects of pinealectomy and crocin treatment in isoproterenol-induced myocardial damage. Seventy rats were divided into seven groups: control, sham control, pinealectomy (PNX), isoproterenol (ISO; 85 mg/kg on the 29th and 30th days of the experiment, subcutaneous injection), PNX + ISO, PNX + crocin (50 mg/kg/day for 30 days, intragastric administration), and PNX + ISO + crocin. PNX procedure was performed on the first day of the study. A significant increase was observed in serum cardiac damage markers (CK-MB, Troponin I) after ISO administration. ISO administration led to a significant increase in cardiac oxidative stress parameters, such as malondialdehyde (MDA) and total oxidant status (TOS), while it led to a decrease in antioxidant defense system parameters, such as reduced glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and total antioxidant status (TAS) when compared to control groups. Elevated MDA and TOS levels were observed, while reduced SOD and CAT activities, and decreased GSH and TAS levels were observed in the group that underwent PNX and ISO administration when compared to the PNX group. Furthermore, in the PNX + ISO + Crocin group, SOD and CAT activities, and GSH and TAS levels ameliorated and MDA and TOS levels were reduced with the crocin treatment when compared to the PNX + ISO group. Also, marked increases were observed in serum cardiac markers, histopathological and immunohistochemical findings after the crocin treatment. All findings demonstrated that crocin could be employed as a cardioprotective agent due to its antioxidant, anti-inflammatory, and anti-apoptotic properties.

The Role of Melatonin on Behavioral Changes and Concomitant Oxidative Stress in icvAß1-42 Rat Model with Pinealectomy.

One of the pathological hallmarks of Alzheimer's disease (AD) associated with its progression that contributes to ß-amyloid (Aß) generation is oxidative stress (OS). Clinical data suggest that melatonin is a potent antioxidant that might be effective in the adjunctive therapy of this neurodegenerative disease. The present study aimed to explore the role of melatonin on behavioral changes and markers of OS in three rat models, namely, pinealectomy (pin) model of melatonin deficit, intracerebroventricular (icv)Aß1-42 model of AD, and combination of both pin and Aß1-42 model (pin+icvAß1-42). The chronic injection with vehicle/melatonin (50 mg/kg, i.p. for 40 days) started on the same day of sham/pin and icv vehicle/Aß1-42 infusion procedures. Anxiety in the open field and the elevated plus-maze test and cognitive responses in the object recognition test were tested between the 30th-35th day after the surgical procedures. Markers of OS in the frontal cortex (FC) and hippocampus were detected by the ELISA method. Melatonin treatment corrected the exacerbated anxiety response only in the pin+icvAß1-42 model while it alleviated the cognitive impairment in the three models. Pinealectomy disturbed the antioxidant system via enhanced SOD activity and decreased GSH levels both in the FC and hippocampus. The Aß1-42 model decreased the SOD activity in the FC and elevated the MDA level in the two brain structures. The pin+icvAß1-42 model impaired the antioxidant system and elevated lipid peroxidation. Melatonin supplementation restored only the elevated MDA level of icvAß1-42 and pin+icvAß1-42 model in the hippocampus. In conclusion, our study reveals that the pin+icvAß1-42 rat model triggers more pronounced anxiety and alterations in markers of OS that may be associated with melatonin deficit concomitant to icvAß1-42-induced AD pathology.

Pinealectomy and gonadectomy modulate amplitude, but not photoperiodic modulation of Clock gene expression in the Syrian hamster suprachiasmatic nuclei.

The duration of daytime light phase (photoperiod) controls reproduction in seasonal mammals. Syrian hamsters are sexually active when exposed to long photoperiod, while gonadal atrophy is observed after exposure to short photoperiod. The photorefractory period, or photorefractoriness, is a particular state of spontaneous recrudescence of sexual activity that occurs after a long-term exposure to short photoperiod. Expression of core clock genes in the master circadian clock contained in the suprachiasmatic nuclei depends on photoperiodic conditions. Interestingly, the expression of the Clock gene is also modified in photorefractory Syrian hamsters. Since melatonin and testosterone levels in seasonal species are dependent on photoperiod, photoperiodic variations of Clock mRNA levels in the suprachiasmatic clock could be a consequence of these hormonal changes. To test this hypothesis, we analysed the effects of pinealectomy on Clock mRNA changes due to long to short photoperiod transition and of gonadectomy on Clock mRNA levels in photorefractory period. Our data show that the suprachiasmatic integration of the short photoperiod (assessed by a rhythmic expression profile of Clock) is independent of the presence of melatonin. Furthermore, constitutively low expression of Clock observed during the photorefractory period does not require the presence of either melatonin or testosterone. However, we show that both hormones provide positive feedback on average levels of Clock expression. Thus, our data support the hypothesis that daily variations of Clock levels in the suprachiasmatic nuclei are influenced by photoperiodic changes and the time spent in short photoperiod, independently of seasonal modifications of melatonin or testosterone levels.

Effect of pinealectomy and melatonin supplementation on metallothionein, ZnT2, ZIP2, ZIP4 and zinc levels in rat small intestine.

We investigated the relations among levels of metallothionein (MT); zinc (Zn) transport proteins, ZnT2, ZIP2 (ZRT and IRT-like proteins); and ZIP4, which enable Zn absorption in the small intestine of rats. We also investigated tissue Zn levels in the small intestine. We used four groups of adult male rats: group 1, control; group 2, pinealectomy (Px); group 3, Px + melatonin (MEL); group 4, MEL only. Animals in groups 3 and 4 were administered 5 mg/kg/day MEL for four weeks. At the end of the study, all animals were sacrificed and samples of duodenum, jejunum and ileum were harvested to analyze ZnT2, ZIP2, ZIP4 and MT levels using immunohistochemistry, and tissue Zn levels were measured by atomic absorption spectrophotometry. The lowest ZnT2 levels in the duodenum, jejunum and ileum, and the lowest ZIP2 levels in the duodenum and ileum were found in group 2. The lowest ZIP4 levels were found in the duodenum and jejunum, and the lowest MT levels in the duodenum and ileum were found in group 2. The highest MT values in the ileum were found in group 4. We found that ZnT2, ZIP2, ZIP4 and MT levels were reduced in the ileum compared to controls following Px, but levels approached control values after MEL administration. By its effects on ZnT2, ZIP2, ZIP4 and MT levels, MEL participates in the absorption of Zn in the rat small intestine.

Pinealectomy increases thermogenesis and decreases lipogenesis.

The present study was designed to determine the effects of pineal gland­derived melatonin on obesity by employing a rat pinealectomy (Pnx) model. After 10 weeks of a high­fat diet, rats received sham or Pnx surgery followed by a normal chow diet for 10 weeks. Reverse transcription­quantitative PCR, western blotting analysis, immunohistochemistry and ELISA were used to determine the effects of Pnx. Pnx decreased the expression of melatonin receptor (MTNR)1A and MTNR1B, in brown adipose tissues (BAT) and white adipose tissues (WAT). Pnx rats showed increased insulin sensitivity compared with those that received sham surgery. Leptin levels were significantly decreased in the serum of the Pnx group. In addition, Pnx stimulated thermogenic genes in BAT and attenuated lipogenic genes in both WAT and the liver. Histological analyses revealed a marked decrease in the size of lipid droplets and increased expression of uncoupling protein 1 in BAT. In the liver of the Pnx group, the size and number of lipid droplets had also decreased. In conclusion, the results presented in the current study suggested that Pnx increases thermogenesis in BAT and decreases lipogenesis in WAT and the liver.

The Influence of Melatonin on the Daily 24-h Rhythm of Putative Reference Gene Expression in White Adipose Tissues.

In adipose tissue, the expression of hundreds of genes exhibits circadian oscillation, which may or may not be affected by circulating melatonin levels. Using control and pinealectomized rats, we investigated the daily expression profile of Actb, Hprt-1, B2m, and Rpl37a, genes that are commonly used as reference genes for reverse transcription quantitative polymerase chain reaction (RT-qPCR), in epididymal (EP), retroperitoneal (RP), and subcutaneous (SC) adipose tissues. In control rats, Actb expression presented a daily oscillation in all adipose tissues investigated, Hprt-1 showed 24-h fluctuations in only RP and SC depots, B2m was stable over 24 h for EP and RP but oscillated over 24 h in SC adipose tissue, and Rpl37a presented a daily oscillation in only RP fat. In the absence of melatonin, the rhythmicity of Actb in all adipose depots was abolished, the daily rhythmicity of Hprt-1 and B2m was disrupted in SC fat, the peak expression of Rpl37a and Hprt-1 was delayed, and the amplitude of Rpl37a was reduced in RP adipose tissue. Collectively, our results demonstrate that the expression of putative reference genes displays a daily rhythm influenced by melatonin levels in a manner specific to the adipose depot. Thus, the proper standardization and daily profile expression of reference genes should be performed carefully in temporal studies using RT-qPCR analysis.

Melatonin regulates oxidative stress and apoptosis in fetal hearts of pinealectomised RUPP rats.

OBJECTIVE: This study aimed to investigate the effects of melatonin on cardiac oxidative stress and apoptosis in the fetal heart in RUPP rats. METHODS: The fetal heart samples were obtained from melatonin administrated RUPP rats. RESULTS: Our results indicate that preeclampsia exacerbated by melatonin deficiency triggers hypoxic conditions, both mis/un-folded protein response, oxidative stress-induced DNA damage and apoptosis. Melatonin treatment provided significant therapeutic effects on fetal hearts via regulating all these stress response at cellular and molecular levels. CONCLUSION: Melatonin may be considered as a potential molecule for development of preventive strategies to reduce the PE induced risk of cardiovascular diseases in offspring.

Endurance training exerts time-dependent modulation on depressive responses and circadian rhythms of corticosterone and BDNF in the rats with pinealectomy.

Pinealectomy can cause a disturbance in emotional status and circadian rhythms of the endocrine and metabolic functions in the body. Endurance training is considered a part of the complex therapy of dysfunctions driven by changes in circadian dynamics of many physiological indicators. In the present study, we aimed to study the effect of endurance training on depressive behavior induced by pinealectomy in rat. We tested the hypothesis that endurance training can have a beneficial impact on depressive behavior induced by pinealectomy in rat via correction of desynchronized circadian rhythms of corticosterone secretion in plasma and brain-derived neurothrophic factor (BDNF) in the hippocampus. The continuous exercise program attenuated depressive responses characterized by the disrupted diurnal rhythm of home-cage motor activity, anhedonia in the sucrose preference test, decreased grooming in the splash test, and despair-like behavior in the forced swimming test of rats with pinealectomy to values resembling those of sham-treated controls. Parallel to the observed positive effect on the emotional status, exercise training diminished total plasma corticosterone levels and corrected its flattened pattern. While the melatonin deficiency did not affect the fluctuations of the BDNF levels, the exercise program induced a considerable and time-dependent increase in its level. These findings suggest that the antidepressant-like effect of endurance training might be mediated via correction of the disturbed circadian rhythm of corticosterone release and enhancement of hippocampal BDNF levels in rats with pinealectomy. Therefore, this alternative mode might have a potential therapeutic application in a subpopulation of people characterized by a melatonin deficiency.

Melatonin decreases cocaine-induced locomotor activity in pinealectomized rats

Objective: Several studies have shown that the time of day regulates the reinforcing effects of cocaine. Additionally, melatonin and its MT1 and MT2 receptors have been found to participate in modulation of the reinforcing effects of such addictive drugs as cocaine. Loss of the diurnal variation in cocaine-induced locomotor sensitization and cocaine-induced place preference has been identified in pinealectomized mice. In addition, several studies in rodents have shown that administration of melatonin decreased the reinforcing effects of cocaine. The objective of this study was to evaluate the effect of melatonin on cocaine-induced locomotor activity in pinealectomized rats at different times of day (zeitgeber time [ZT]4, ZT10, ZT16, and ZT22). Methods: Naïve, pinealectomized Wistar rats received cocaine at different times of day. Melatonin was administered 30 min before cocaine; luzindole was administered 15 min prior to melatonin and 45 min before cocaine. After administration of each treatment, locomotor activity for each animal was recorded for a total of 30 min. Pinealectomy was confirmed at the end of the experiment through melatonin quantitation by ELISA. Results: Cocaine-induced locomotor activity varied according to the time of day. Continuous lighting and pinealectomy increased cocaine-induced locomotor activity. Melatonin administration decreased cocaine-induced locomotor activity in naïve and pinealectomized rats at different times of day. Luzindole blocked the melatonin-induced reduction in cocaine-induced locomotor activity in pinealectomized rats. Conclusion: Given its ability to mitigate various reinforcing effects of cocaine, melatonin could be a useful therapy for cocaine abuse.

Evidence that Melatonin Increases Inhibin Beta-A and Follistatin Gene Expression in Ovaries of Pinealectomized Rats.

Melatonin plays an important role in the regulation of ovarian function including oocyte maturation in different mammalian species. Many studies indicate that melatonin has an impact on the ovarian function of a variety of ovarian cells. However, the information on the exact mechanism and involved hormones is low. To evaluate inhibin beta-A (INHBA) and follistatin (FST) expression in the ovaries of pinealectomized rats treated with melatonin, thirty adult female Wistar rats were randomized into three groups of ten animals each: group 1 (GSh), sham-operated controls receiving vehicle; group 2 (GPx), pinealectomized animals receiving vehicle; and group 3 (GPxMe), pinealectomized animals receiving replacement melatonin (1.0 mg/kg body weight. It was assumed that each animal drank 6.5 ± 1.2 ml per night and weighs approximately 300 g.) for 60 consecutive days. The ovaries were collected for mRNA abundance and protein of INHBA and FST by qRT-PCR and immunohistochemical analyses, respectively. Treatment with melatonin resulted in the upregulation of INHBA and FST genes in the ovarian tissue of the melatonin-treated animals (GPxMe), when compared with GPx. These findings were then confirmed by analyzing the expression of protein by immunohistochemical analyses, which revealed higher immunoreactivity of INHBA and FST in GPxMe animals in the follicular cells compared with GSh and GPx rats. Melatonin increases the expression of INHBA and FST in the ovaries of pinealectomized female rats.

Effect of melatonin on T/B cell activation and immune regulation in pinealectomy mice.

Melatonin is an indole neuroendocrine hormone that is mainly secreted by the pineal gland to regulate circadian rhythm, antioxidation, and immune regulation. Melatonin plays an important role in T cell-mediated immune responses against cancer, infections, and the development of many autoimmune diseases. The aim of this study was to investigate the immunomodulatory effects of melatonin on T/B cell activation in pinealectomy mice. The improved pinealectomy procedure for mice presented in this study is a good animal model to be used in follow-up studies on melatonin. After pinealectomy, the tissue removed was identified as the pineal body using HE staining. The effects of melatonin supplementation on T cell activation and activation-related changes to the MAPK/NF-κ B pathways were analyzed by flow cytometry and real-time PCR. We found that expression levels of Th1, Th2 and Th17-related cytokines in peripheral blood were lower in mice that had undergone pinealectomy, compared with normal mice. After melatonin supplementation, cytokine levels rapidly increased within a short period of time, which resulted in the gradual recovery of cytokine expression levels. Moreover, activation of T/B cells in mice was weakened and decreased after pineal gland removal. Melatonin was found to inhibit the expression of TLR3, p38, JNK, and MAPK/NF-κ B within a short period (2 weeks) of melatonin replenishment. This inhibition gradually weakened with time, since the degree of inhibition is negatively related with the dosage of melatonin. In conclusion, melatonin may regulate the activation of T/B cells, playing a critical role in the regulation of immune balance.