Search

Yaseen Awad-Igbaria,1,2 Shilo Dadon,1,2 Alon Shamir,3,4 Alejandro Livoff,5 Mark Shlapobersky,5 Jacob Bornstein,1,2,* Eilam Palzur2,* 1Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel; 2The Research Institute of Galilee Medical Center, Nahariya, Israel; 3Psychobiology Research Laboratory, Mazor Mental Health Center, Akko, Israel; 4The Ruth and Bruce Rappaport Faculty of Medicine, Technion – Israel Institute of Technology, Haifa, Israel; 5Pathology Department, Barzilai University Medical Center, Ashkelon, Israel*These authors contributed equally to this workCorrespondence: Yaseen Awad-Igbaria, The Research Institute of Galilee Medical Center, PO Box 21, Nahariya, 22100, Israel, Tel +972 50-4500101, Email Yaseenawad123@gmail.comBackground: Provoked vulvodynia (PV) is the main cause of vulvar pain and dyspareunia. The etiology of PV has not yet been elucidated. However, PV is associated with a history of recurrent inflammation, and its often accompanied by increases in the numbers of mast cells (MCs) and sensory hyperinnervation in the vulva. Therefore, this study aimed to examine the role of MCs and the early inflammatory events in the development of chronic vulvar pain in a rat model of PV.Methods: Mechanical and thermal vulvar sensitivity was measured for 5 months following zymosan vulvar challenges. Vulvar changes in glutamate and nerve growth factor (NGF) were analyzed using ELISA. Immunofluorescence (IF) staining of the vulvar section after 20, 81, and 160 days of the zymosan challenge were performed to test MCs accumulation, hyperinnervation, and expression of pain channels (transient receptor potential vanilloid/ankyrin-1-TRPV1 & TRPA1) in vulvar neurons. Changes in the development of vulvar pain were evaluated following the administration of the MCs stabilizer ketotifen fumarate (KF) during zymosan vulvar challenges.Results: Zymosan-challenged rats developed significant mechanical and thermal vulvar sensitivity that persisted for over 160 days after the zymosan challenge. During inflammation, increased local concentrations of NGF and glutamate and a robust increase in MCs degranulation were observed in zymosan-challenged rats. In addition, zymosan-challenged rats displayed sensory hyperinnervation and an increase in the expression of TRPV1 and TRPA1. Treatment with KF attenuated the upregulated level of NGF during inflammation, modulated the neuronal modifications, reduced MCs accumulation, and enhanced mechanical hypersensitivity after repeated inflammation challenges.Conclusion: The present findings suggest that vulvar hypersensitivity is mediated by MCs accumulation, nerve growth, and neuromodulation of TRPV1 and TRPA1. Hence, KF treatment during the critical period of inflammation contributes to preventing chronic vulvar pain development.Keywords: provoked vulvodynia, inflammation, mast cell, nerve growth factor, glutamate, hyperinnervation

Provoked vulvodynia (PV) is the leading cause of vulvar pain and dyspareunia. The etiology of PV is multifactorial and remains poorly understood. PV is associated with a history of repeated vulvar inflammation and is often accompanied by sensory neuromodulation as a result of activation of the metabotropic glutamate receptor 5 (mGluR5) in the sensory nerve terminals. Therefore, this study aims to examine the role of glutamate-mGluR5 signaling during the initial inflammatory phase in chronic vulvar pain development in an animal model of PV.Thermal and mechanical vulvar sensitivity was assessed for three weeks following zymosan vulvar challenges. Anxiety-like behavior and locomotor activity were assessed at the end of the experiment. To investigate the role of glutamate mGluR5, the MTEP (mGluR5 antagonist) was injected into the vulva during vulvar inflammation. On the other hand, glutamate or CHPG (mGluR5 agonist) were injected in order to examine the effects of mGluR5 activation. RT-PCR was performed to assess changes in the transcription of genes related to neuroinflammation, neuromodulation, and neuroplasticity in the spinal cord (L6-S3). Zymosan-induced inflammation resulted in a significant thermal and mechanical vulvar hypersensitivity that persisted for over a month after the zymosan injection. However, local treatment with MTEP enhanced the vulvar mechanical and thermal hypersensitivity. On the other hand, activation of the mGluR5 via injection of glutamate or CHPG into the vulva leads to long-lasting vulvar mechanical and thermal hypersensitivity. The activation of the glutamate pathway was found to be accompanied by an increase in the transcription level of genes related to neuroinflammation and neuroplasticity in the sacral spine region. The present findings indicate that vulvar hypersensitivity is mediated by mGluR5 activation during inflammation. Hence, modulation of the mGluR5 pathway during the critical period of inflammation contributes to preventing chronic vulvar pain development. Conversely, activation of the mGluR5 pathway leads to long-lasting mechanical and thermal hypersensitivity.

Introduction: Hyperbaric oxygen has been used as a medical treatment tool in hyperbaric chambers and is an integral part of professional and combat divers' activity. In extreme cases, exposure to hyperbaric oxygen can develop central nervous system oxygen toxicity (CNS-OT), which leads to seizures and eventually death. CNS-OT is caused by neuronal hyperactivity due to high oxygen levels, potentially damaging brain cells including the blood-brain barrier (BBB). However, the effect of hyperbaric oxygen levels on the healthy BBB has not been characterized directly yet., Methods: Six or three different groups of ~ eight rats or mice, respectively, were exposed to increasing levels of partial pressure of oxygen (0.21 to 5 ATA) in a hyperbaric chamber, followed by MRI scanning with gadolinium. Statistical significance (adjusted p-value ≤ 0.05) was assessed using linear regression and ordinary one-way (rats) or two-way (mice) ANOVA with correction of multiple comparison tests. In rats, the effect of 100% oxygen at 5 ATA was independently validated using FITC-Dextran (5 kDa). Statistical significance (p-value ≤ 0.05) was assessed using Welch's t-test and effect size was calculated by Cohen's D., Results: In rats, analyzed MRI scans showed a significant trend of increase in the % gadolinium in brain tissues as a result of hyperbaric oxygen pressures (p-value = 0.0079). The most significant increase was measured at 4 ATA compared to air (adjusted p-value = 0.0461). Significant increased FITC-Dextran levels were measured in the rats' brains under 100% oxygen at 5 ATA versus air (p-value = 0.0327; Effect size = 2.0). In mice, a significant increase in gadolinium penetration into the hippocampus and frontal cortex was measured over time (adjusted p-value < 0.05) under 100% oxygen at 3 and 5 ATA versus air, and between the treatments (adjusted p-value < 0.0001)., Conclusions: The BBB is increasingly disrupted due to higher levels of hyperbaric oxygen in rodents, indicating a direct relation between hyperbaric oxygen and BBB dysregulation for the first time. We suggest considering this risk in different diving activities, and protocols using a hyperbaric chamber. On the other hand, this study highlights the potential therapeutic usage of hyperbaric oxygen for controlled drug delivery through the BBB into brain tissues in different brain-related diseases., (© 2024. The Author(s).)

Provoked vulvodynia represents a challenging chronic pain condition, characterized by its multifactorial origins. The inherent complexities of human-based studies have necessitated the use of animal models to enrich our understanding of vulvodynia's pathophysiology. This review aims to provide an exhaustive examination of the various animal models employed in this research domain. A comprehensive search was conducted on PubMed, utilizing keywords such as "vulvodynia", "chronic vulvar pain", "vulvodynia induction", and "animal models of vulvodynia" to identify pertinent studies. The search yielded three primary animal models for vulvodynia: inflammation-induced, allergy-induced, and hormone-induced. Additionally, six agents capable of triggering the condition through diverse pathways were identified, including factors contributing to hyperinnervation, mast cell proliferation, involvement of other immune cells, inflammatory cytokines, and neurotransmitters. This review systematically outlines the various animal models developed to study the pathogenesis of provoked vulvodynia. Understanding these models is crucial for the exploration of preventative measures, the development of novel treatments, and the overall advancement of research within the field.

Traumatic brain injury (TBI) is a leading cause of morbidity and mortality in young adults, characterized by primary and secondary injury. Primary injury is the immediate mechanical damage, while secondary injury results from delayed neuronal death, often linked to mitochondrial damage accumulation. Hyperbaric oxygen therapy (HBOT) has been proposed as a potential treatment for modulating secondary post-traumatic neuronal death. However, the specific molecular mechanism by which HBOT modulates secondary brain damage through mitochondrial protection remains unclear. Spatial learning, reference memory, and motor performance were measured in rats before and after Controlled Cortical Impact (CCI) injury. The HBOT (2.5 ATA) was performed 4 h following the CCI and twice daily (12 h intervals) for four consecutive days. Mitochondrial functions were assessed via high-resolution respirometry on day 5 following CCI. Moreover, IHC was performed at the end of the experiment to evaluate cortical apoptosis, neuronal survival, and glial activation. The current result indicates that HBOT exhibits a multi-level neuroprotective effect. Thus, we found that HBOT prevents cortical neuronal loss, reduces the apoptosis marker (cleaved-Caspase3), and modulates glial cell proliferation. Furthermore, HBO treatment prevents the reduction in mitochondrial respiration, including non-phosphorylation state, oxidative phosphorylation, and electron transfer capacity. Additionally, a superior motor and spatial learning performance level was observed in the CCI group treated with HBO compared to the CCI group. In conclusion, our findings demonstrate that HBOT during the critical period following the TBI improves cognitive and motor damage via regulating glial proliferation apoptosis and protecting mitochondrial function, consequently preventing cortex neuronal loss.

Background: Peripheral nerve injury can cause neuroinflammation and neuromodulation that lead to mitochondrial dysfunction and neuronal apoptosis in the dorsal root ganglion (DRG) and spinal cord, contributing to neuropathic pain and motor dysfunction. Hyperbaric oxygen therapy (HBOT) has been suggested as a potential therapeutic tool for neuropathic pain and nerve injury. However, the specific cellular and molecular mechanism by which HBOT modulates the development of neuropathic pain and motor dysfunction through mitochondrial protection is still unclear., Methods: Mechanical and thermal allodynia and motor function were measured in rats following sciatic nerve crush (SNC). The HBO treatment (2.5 ATA) was performed 4 h after SNC and twice daily (12 h intervals) for seven consecutive days. To assess mitochondrial function in the spinal cord (L2-L6), high-resolution respirometry was measured on day 7 using the OROBOROS-O2k. In addition, RT-PCR and Immunohistochemistry were performed at the end of the experiment to assess neuroinflammation, neuromodulation, and apoptosis in the DRG (L3-L6) and spinal cord (L2-L6)., Results: HBOT during the early phase of the SNC alleviates mechanical and thermal hypersensitivity and motor dysfunction. Moreover, HBOT modulates neuroinflammation, neuromodulation, mitochondrial stress, and apoptosis in the DRG and spinal cord. Thus, we found a significant reduction in the presence of macrophages/microglia and MMP-9 expression, as well as the transcription of pro-inflammatory cytokines (TNFa, IL-6, IL-1b) in the DRG and (IL6) in the spinal cord of the SNC group that was treated with HBOT compared to the untreated group. Notable, the overexpression of the TRPV1 channel, which has a high Ca 2+ permeability, was reduced along with the apoptosis marker (cleaved-Caspase3) and mitochondrial stress marker (TSPO) in the DRG and spinal cord of the HBOT group. Additionally, HBOT prevents the reduction in mitochondrial respiration, including non-phosphorylation state, ATP-linked respiration, and maximal mitochondrial respiration in the spinal cord after SNC., Conclusion: Mitochondrial dysfunction in peripheral neuropathic pain was found to be mediated by neuroinflammation and neuromodulation. Strikingly, our findings indicate that HBOT during the critical period of the nerve injury modulates the transition from acute to chronic pain via reducing neuroinflammation and protecting mitochondrial function, consequently preventing neuronal apoptosis in the DRG and spinal cord., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: Substantial evidence suggests that hypertension is a significant risk factor for cognitive decline. However, it is unclear whether the genetic predisposition to hypertension is also associated with cellular dysfunction that promotes neurodegeneration., Methods: Changes in blood pressure were evaluated following dietary salt-loading or administration of a regular diet in Sabra Normotensive (SBN/y) and Sabra Hypertension-prone rats (SBH/y). We performed quantitative RT-PCR and immunofluorescence staining in brain cortical tissues before salt loading and 6 and 9 months after salt loading. To examine the expression of brain cortical proteins involved in the gene regulation (Histone Deacetylase-HDAC2; Histone Acetyltransferase 1-HAT1), stress response (Activating Transcription Factor 4-ATF4; Eukaryotic Initiation Factor 2- eIF2α), autophagy (Autophagy related 4A cysteine peptidase- Atg4a; light-chain 3-LC3A/B; mammalian target of rapamycin complex 1- mTORC1) and apoptosis (caspase-3)., Results: Prior to salt loading, SBH/y compared to SBN/y expressed a significantly higher level of cortical HAT1 (protein), Caspase-3 (mRNA/protein), LC3A, and ATF4 (mRNA), lower levels of ATG4A (mRNA/protein), LC3A/B, HDAC2 (protein), as well as a lower density of cortical neurons. Following dietary salt loading, SBH/y but not SBN/y developed high blood pressure. In hypertensive SBH/y, there was significant upregulation of cortical HAT1 (protein), Caspase-3 (protein), and eIF2α ~ P (protein) and downregulation of HDAC2 (protein) and mTORC1 (mRNA), and cortical neuronal loss., Conclusions: The present findings suggest that genetic predisposition to hypertension is associated in the brain cortex with disruption in autophagy, gene regulation, an abnormal response to cellular stress, and a high level of cortical apoptosis, and could therefore exacerbate cellular dysfunction and thereby promote neurodegeneration., (© 2023. The Author(s).)

This short opinion aimed to present the evidence to support our hypothesis that vulvodynia is a neuroinflammatory pain syndrome originating in the pelvic visceral nerve plexuses caused by the failure of weakened uterosacral ligaments (USLs) to support the pelvic visceral nerve plexuses, i.e., T11-L2 sympathetic and S2-4 parasympathetic plexuses. These are supported by the USLs, 2 cm from their insertion to the cervix. They innervate the pelvic organs, glands, and muscles. If the USLs are weak or lax, gravitational force or even the muscles may distort and stimulate the unsupported plexuses. Inappropriate afferent signals could then be interpreted as originating from an end-organ site. Activation of sensory visceral nerves causes a neuro-inflammatory response in the affected tissues, leading to neuroproliferation of small peripheral sensory nerve fibers, which may cause hyperalgesia and allodynia in the territory of the damaged innervation. Repair of the primary abnormality of USL laxity, responsible for mechanical stimulation of the pelvic sensory plexus, may lead to resolution of the pain syndrome., (© 2022. The Author(s).)

The opening of the mitochondrial permeability transition pore (mPTP) has emerged as a pivotal event following traumatic brain injury (TBI). Evidence showing the impact of the translocator protein (TSPO) over mPTP activity has prompted several studies exploring the effect of TSPO ligands, including etifoxine, on the outcome of traumatic brain injury (TBI). Mitochondrial respiration was assessed by respirometry in isolated rat brain mitochondria (RBM) by measurements of oxidative phosphorylation capacity (OXPHOS). The addition of calcium to RBM was used to induce mitochondrial injury and resulted in significant OXPHOS reduction that could be reversed by preincubation of RBM with etifoxine. Sensorimotor and cognitive functions were assessed following controlled cortical impact and compared in vehicle and etifoxine-treated animals. There was no difference between the vehicle and etifoxine groups for sensorimotor functions as assessed by rotarod. In contrast, etifoxine resulted in a significant improvement of cognitive functions expressed by faster recovery in Morris water maze testing. The present findings show a significant neuroprotective effect of etifoxine in TBI through restoration of oxidative phosphorylation capacity associated with improved behavioral and cognitive outcomes. Since etifoxine is a registered drug used in common clinical practice, implementation in a phase II study may represent a reasonable step forward.

Background: Serum biochemical changes during laparoscopic surgery and positive pressure pneumoperitoneum (PP) may reflect mild oxidative stress due to the ischemia-reperfusion (I/R) mechanism. However, there is still a controversy regarding the exact mechanism of PP in creating oxidative stress and whether the induction of PP causes I/R effects at all. To elucidate this debated issue, we studied, for the first time, the changes of I/R parameters in the serum, in a pilot study, during laparoscopic cholecystectomy using a reliable, independent exogenous oxidative biomarker, together with common intrinsic biomarkers of oxidative stress., Patients and Methods: Our study included 20 patients scheduled for elective laparoscopic cholecystectomy. We evaluated the levels of the extrinsic and endogenous markers for oxidative stress during awareness, under anesthesia, the end of surgery (abdominal CO2 evacuation), and 2 h afterward., Results: After an initial increase in oxidative stress following anesthesia, we did not notice any further significant rise in the levels of the synthetic exogenous and the endogenous biomarkers at the end of the surgery and 2 h later on. However, a positive correlation was noted between the levels of both the intrinsic and extrinsic markers., Conclusions: In our study, the capability of the extrinsic biomarker to detect mild oxidative stress was not validated. Our study stresses the heterogeneous nature of the oxidative reactions and the diversity of the endogenous and exogenous biomarkers while detecting various biochemical patterns under mild oxidative stress, during the short period of laparoscopic surgery., (© 2021. The Author(s).)

Experimental studies have shown that ligands of the 18 kDa translocator protein can reduce neuronal damage induced by traumatic brain injury by protecting mitochondria and preventing metabolic crisis. Etifoxine, an anxiolytic drug and 18 kDa translocator protein ligand, has shown beneficial effects in the models of peripheral nerve neuropathy. The present study investigates the potential effect of etifoxine as a neuroprotective agent in traumatic brain injury (TBI). For this purpose, the effect of etifoxine on lesion volume and modified neurological severity score at 4 weeks was tested in Sprague-Dawley adult male rats submitted to cortical impact contusion. Effects of etifoxine treatment on neuronal survival and apoptosis were also assessed by immune stains in the perilesional area. Etifoxine induced a significant reduction in the lesion volume compared to nontreated animals in a dose-dependent fashion with a similar effect on neurological outcome at four weeks that correlated with enhanced neuron survival and reduced apoptotic activity. These results are consistent with the neuroprotective effect of etifoxine in TBI that may justify further translational research.

Cerebral edema represents a major threat following traumatic brain injury. However, therapeutic measures for control of intracranial pressure alone have failed to restore cerebral metabolism and improve neurological outcome. Since mitochondrial damage results in ATP depletion and deactivation of membrane ionic pumps, we hypothesized that modulation of ATP bioavailability may directly affect cytotoxic edema. Intracranial pressure measurements were performed in Sprague-Dawley rats treated by intraperitoneal injection of dimethylsulfoxide (vehicle), cyclosporine A (CsA), or Oligomycin B (OligB) following cortical contusion and further correlated with water content, mitochondrial damage, and electron microscopic assessment of neuronal and axonal edema. As hypothesized, ultra-structural figures of edema closely correlated with intracranial pressure elevation, increased water content and mitochondrial membrane permeabilization expressed by loss of transmembrane mitochondrial potential. Further, mitochondrial damage evidenced ultra-structurally by figures of swollen mitochondria with severely distorted cristae correlated with both cytotoxic edema and mitochondrial dysfunction. Importantly, cerebral edema and mitochondrial impairment were significantly worsened by treatment with OligB, whereas a noticeable improvement could be observed in animals that received injections of CsA. Since OligB and CsA are responsible for symmetrical and opposite effects on oxidative metabolism, these findings support the hypothesis of a causative relationship between edema and mitochondrial function., (© The Author(s) 2015.)

Traumatic brain injury (TBI) is a leading cause of death and functional disability in western countries, affecting mostly young patients. Despite intense and sustained efforts deployed for the development of new therapeutic strategies, no clinical benefit has been shown by any of the investigated compounds. Increasing attention has been drawn during the past two decades to the neuroprotective effects of estrogens, although most of the available data relate to ischemic brain injury. The purpose of the present study was to investigate the potential neuroprotective value of estrogens in TBI as a therapeutic modality. For this purpose, a contusion was created in the parietal cortex by dynamic cortical deformation in two groups of 10 Sprague-Dawley male rats. Following the injury, treated animals received conjugated estrogens for 3 days, using a subcutaneously implanted osmotic pump. Animals were then sacrificed, and TUNEL, anti-active Caspase 3, bcl-2, and bax labeling were performed in paraffin-embedded brain sections, allowing for comparative and quantitative analysis. In estrogen-treated animals, there was a marked and significant reduction of apoptosis in comparison with non-treated animals. The reduction in TUNEL and active Caspase 3 staining was similar and close to 50%. Optical analysis of histological slides prepared by bcl-2 labeling showed a significant increase in bcl-2 expression in estrogen-treated animals compared to non-treated animals. On the contrary, bax expression was not influenced by hormonal treatment, and no difference could be noticed between the two groups. These results support the potential therapeutic value of estrogens in TBI and further clarify their mode of action.

Cerebral contusions are one the most frequent traumatic lesions and the most common indication for secondary surgical decompression. The purpose of this study was to investigate the physiology of perilesional secondary brain damage and evaluate the value of hyperbaric oxygen therapy (HBOT) in the treatment of these lesions. Five groups of five Sprague-Dawley rats each were submitted to dynamic cortical deformation (DCD) induced by negative pressure applied to the cortex. Cerebral lesions produced by DCD at the vacuum site proved to be reproducible. The study protocol entailed the following: (1) DCD alone, (2) DCD and HBOT, (3) DCD and post-operative hypoxia and HBOT, (4) DCD, post-operative hypoxia and HBOT, and (5) DCD and normobaric hyperoxia. Animals were sacrificed after 4 days. Histological sections showed localized gross tissue loss in the cortex at injury site, along with hemorrhage. In all cases, the severity of secondary brain damage was assessed by counting the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and caspase 3-positive cells in successive perilesional layers, each 0.5 mm thick. Perilesional TUNEL positive cells suggested the involvement of apoptosis in group 1 (12.24% of positive cells in layer 1). These findings were significantly enhanced by post-operative hypoxia (31.75%, p < 0.001). HBOT significantly reduced the severity and extent of secondary brain damage expressed by the number of TUNEL positive cells in each layer and the volume of the lesion (4.7% and 9% of TUNEL positive cells in layer 1 in groups 2 and 4 respectively, p < 0.0001 and p < 0.003). Normobaric hyperoxia also proved to be beneficial although in a lesser extent. This study demonstrates that the vacuum model of brain injury is a reproducible model of cerebral contusion. The current findings also suggest that HBOT may limit the growth of cerebral contusions and justify further experimental studies.