Regenerative Medicine for Polycystic Ovary Syndrome: Stem Cell-Based Therapies and Brown Adipose Tissue Activation.

Polycystic ovary syndrome (PCOS) is a pathological condition prevalent among women of reproductive age: it is associated with varied etiological factors (lifestyle, genetic, environmental…) and characterized by an increased polycystic morphology of the ovaries leading to disturbances in the menstrual cycle and its correlated infertility. Interconnections between PCOS, obesity, and insulin resistance have been recently investigated thoroughly in the scientific community; these findings directed PCOS therapies into unraveling possibilities to target insulin resistance and central adiposity as efficient treatment. On the other hand, brown adipose tissue is known to possess a thermogenic activity that increases lipolysis and directly attenuates fat deposition. Therefore, brown adipose tissue activation lands itself as a potential target for reducing obesity and its induced insulin resistance, subsequently rescuing PCOS phenotypes. In addition, regenerative medicine has proven efficacy in resolving PCOS-associated infertility and its metabolic symptoms. In particular, many stem/progenitor cells have been verified to possess the differentiation capacity into functional brown adipocytes. Thus, throughout this review, we will discuss the different brown adipose tissue activation strategies and stem-cell-based therapies applied to PCOS models and the possible combination of both therapeutic approaches to synergistically act on the activation of brown adipose tissue and attenuate PCOS-correlated infertility and retract the consequences of the metabolic syndrome on the physiological state of patients.

Saffron Extract Attenuates Anxiogenic Effect and Improves Cognitive Behavior in an Adult Zebrafish Model of Traumatic Brain Injury.

Traumatic brain injury (TBI) has the highest mortality rates worldwide, yet effective treatment remains unavailable. TBI causes inflammatory responses, endoplasmic reticulum stress, disruption of the blood-brain barrier and neurodegeneration that lead to loss of cognition, memory and motor skills. Saffron (Crocus sativus L.) is known for its anti-inflammatory and neuroprotective effects, which makes it a potential candidate for TBI treatment. Zebrafish (Danio rerio) shares a high degree of genetic homology and cell signaling pathways with mammals. Its active neuro-regenerative function makes it an excellent model organism for TBI therapeutic drug identification. The objective of this study was to assess the effect of saffron administration to a TBI zebrafish model by investigating behavioral outcomes such as anxiety, fear and memory skills using a series of behavioral tests. Saffron exhibited anxiolytic effect on anxiety-like behaviors, and showed prevention of fear inhibition observed after TBI. It improved learning and enhanced memory performance. These results suggest that saffron could be a novel therapeutic enhancer for neural repair and regeneration of networks post-TBI.

Myostatin gene inactivation increases post-mortem calpain-dependent muscle proteolysis in mice.

Myostatin deficiency leads to extensive skeletal muscle hypertrophy, but its consequence on post-mortem muscle proteolysis is unknown. Here, we compared muscle myofibrillar protein degradation, and autophagy, ubiquitin-proteasome and Ca2+-dependent proteolysis relative to the energetic and redox status in wild-type (WT) and myostatin knock-out mice (KO) during early post-mortem storage. KO muscles showed higher degradation of myofibrillar proteins in the first 24 h after death, associated with preserved antioxidant status, compared with WT muscles. Analysis of key autophagy and ubiquitin-proteasome system markers indicated that these two pathways were not upregulated in post-mortem muscle (both genotypes), but basal autophagic flux and ATP content were lower in KO muscles. Proteasome and caspase activities were not different between WT and KO mice. Conversely, calpain activity was higher in KO muscles, concomitantly with higher troponin T and desmin degradation. Altogether, these results suggest that calpains but not the autophagy, proteasome and caspase systems, explain the difference in post-mortem muscle protein proteolysis between both genotypes.

Addressing the Importance of Stem Cell-Based Therapy: A Perspective in the Treatment of COVID-19.

Severe acute respiratory syndrome-associated coronavirus 2 (SARS-CoV-2) is an extremely pathogenic virus belonging to the family of Coronaviridae. First identified in Wuhan, China in December 2019 after an epidemiological investigation of an emerging cluster of pneumonia of unknown etiology, SARS-CoV-2 was declared the cause of a pandemic on March 11 by the World Health Organization (WHO), pointing to the over 118000 cases of Coronavirus disease 2019 (COVID-19) in over 110 countries. Despite the promising results of drug repositioning studies in the treatment of COVID-19, the evidence of their safety and efficacy remains inconclusive. Cell based therapy has been proven safe and possibly effective in treating multiple lung injuries and diseases, but its potential use in the treatment of COVID-19 has not been yet elucidated. Our aim in this review is to provide an overview of the immunomodulatory effect and the regenerative capacity of stem cells and their secretome in the treatment of many diseases including lung injuries. Those findings may contribute to a better understanding of the potential of stem cell therapy in SARS-CoV-2 infection and its potential use in order to find a solution for this healthcare crisis.

Zebrafish toxicity assessment of the photocatalysis-biodegradation of diclofenac using composites of TiO2 and activated carbon from Argania spinosa tree nutshells and Pseudomonas aeruginosa.

The occurrence and persistence of pharmaceutical products (PPs) in the environment have recently been well-documented and are a major concern for public health. Their incidence in aquatic ecosystems is the result of their direct release without any prior treatment or insufficient wastewater treatment. Therefore, an efficient and safe posttreatment process for removing PPs must be developed. In this study, we focused on the ability of photocatalysis or combined photocatalysis and biodegradation to effectively and safely remove diclofenac (DCF) and its by-products from water. The heterogeneous photocatalysis system was based on bio-sourced activated carbon obtained from Argania spinosa tree nutshells and Degussa P25 titanium dioxide (ACP-TiO2), and biodegradation involved Pseudomonas aeruginosa. Toxicity tests were conducted with zebrafish embryos to evaluate the applicability of the treatment processes. The results showed that photocatalytic treatment with 0.1 mg/L of ACP-TiO2 9% for 7.5 h is sufficient to eliminate DCF (50 mg L-1) and its by-products from water. Low levels of malformation (< 20%) were detected in zebrafish embryos treated with photocatalyzed DCF solutions at 1, 5, and 7 mg L-1 after 4 days of exposure. After 3 h of incubation, P. aeruginosa was found to reduce the toxicity of DCF (10 mg L-1) photocatalyzed for 2 and 4 h. Additional studies should be conducted to elucidate the biodegradation mechanism.

Ferula hermonis: A Review of Current Use and Pharmacological Studies of its Sesquiterpene Ester Ferutinin.

Ferula hermonis Boiss, is an endemic plant of Lebanon, locally known as "shilsh Elzallouh". It has been extensively used in the traditional medicine as an aphrodisiac and for the treatment of sexual impotence. Crude extracts and isolated compounds of ferula hermonis contain phytoestrogenic substances having a wide spectrum of in vitro and in vivo pharmacological properties including anti-osteoporosis, anti-inflammatory, anti-microbial and anti-fungal, anti-cancer and as sexual activity enhancer. The aim of this mini-review is to highlight the traditional and novel applications of this plant's extracts and its major sesquiterpene ester, ferutinin. The phytochemical constituents and the pharmacological uses of ferula hermonis crude extract and ferutinin specifically will be discussed.

P-glycoprotein modulates oleanolic acid effects in hepatocytes cancer cells and zebrafish embryos.

Oleanolic acid (OA) is a triterpenoid, widely found in plants and possesses antitumor activity in many cancer lines. However, cancer cells develop multidrug resistance (mdr) hindering the effect of anticancer drugs. P-glycoprotein (P-gp) is a major cause of mdr. Therefore, the cytotoxic effect of OA was evaluated on human breast cancer MDA-MB-231 and human liver cancer HepG2 with absence and presence of P-gp, respectively. OA reduced MDA-MB-231 viability in a dose dependent manner, whereas no remarkable effect was observed on HepG2 in the same range of concentrations (1-60 µM). Moreover, cytotoxicity studies were conducted in the presence of verapamil (20 mg/L), a P-gp inhibitor. OA exhibited the same effect on MDA-MB-231 in the absence and presence of verapamil. However, the cytotoxicity was greatly enhanced for HepG2 cells in the presence of verapamil (cell viability dropped from 63.7% to 25% after 72 h at 60 µM). The results were then confirmed in vivo on zebrafish embryos. Increased mortality and malformations were observed in verapamil pretreated group between 5 and 15 µM of OA compared to control; also, all embryos died at 20 µΜ OA and above. These results demonstrate that inhibiting P-gp enhances the chemotherapeutic activity of OA.

Chemical Composition, Antioxidant and Anti-inflammatory Activity Evaluation of the Lebanese Propolis Extract.

BACKGROUND: Propolis is a resinous substance produced by bees and known to possess antioxidant, antimicrobial, antiproliferative and anti-inflammatory activities. OBJECTIVE: This study is aimed at evaluating the in vivo and in vitro anti-inflammatory potential of the Crude Ethanolic Extract (CE) of Lebanese propolis and its Ethyl Acetate Fraction (EAF). METHOD: Chemical content of propolis was characterized using high-performance liquid chromatography and LC-MS/MS. COX-2 and iNOS protein expression, nitric oxide (NO) and prostaglandin (PGE2) release in LPS-activated RAW monocytes were achieved respectively by western blot and spectrophotometry. Antioxidant activity was evaluated by DPPH free radical scavenging assay. Measurement of paw thickness in carrageenan-induced paw edema in mice and pathologic assessment of inflammation in paw sections were used to judge the anti-inflammatory properties of propolis. RESULTS: Pathology analysis revealed in the treated group significant reduction of immune cell infiltration and edema. Both extract and ethyl acetate fraction showed significant anti-inflammatory and antioxidant effects in LPS-treated RAW cells characterized by the inhibition of COX-2 and iNOS protein expression, as well as PGE2 and NO release. Chemical analysis of the crude extract and its ethyl acetate fraction identified 28 different compounds of which two phenolic acids and nine other flavonoids were also quantified. Ferulic acid, caffeic acid, chrysin, galangin, quercetin, and pinocembrin were among the most representative compounds. CONCLUSION: Lebanese propolis is rich in a various amount of flavonoids which showed promising antiinflammatory and antioxidant properties. Additionally, chemical analysis showed unique chemical compositions with the potential of identifying ingredients with interesting anti-inflammatory activities.

A ferutinin analogue with enhanced potency and selectivity against ER-positive breast cancer cells in vitro.

Estrogen is considered a risk factor for breast cancer since it promotes breast-cell proliferation. The jaesckeanadiol-3-p-hydroxyphenylpropanoate, a hemi-synthetic analogue of the natural phytoestrogen ferutinin (jaesckeanadiol-p-hydroxybenzoate), is designed to be devoid of estrogenic activity. This analogue induces a cytotoxic effect 30 times higher than that of ferutinin towards MCF-7 breast cancer cell line. We compared these two compounds with respect to their effect on proliferation, cell cycle distribution and cancer stem-like cells in the MCF-7 cell line. Treatment with ferutinin (30 µM) and its analogue (1 µM) produced significant accumulation of cells at the pre G0/G1 cell cycle phase and triggered apoptosis. Importantly, this compound retains its anti-proliferative activity against breast cancer stem/progenitor cells that are naturally insensitive to ferutinin at the same dose. These results position ferutinin analogue as an effective compound inhibiting the proliferation of estrogen-dependent breast cancer cells and consistently targeting their stem-like cells.

HIV-1 Tat protein induces DNA damage in human peripheral blood B-lymphocytes via mitochondrial ROS production.

Human immunodeficiency virus (HIV) infection is associated with B-cell malignancies in patients though HIV-1 is not able to infect B-cells. The rate of B-cell lymphomas in HIV-infected individuals remains high even under the combined antiretroviral therapy (cART) that reconstitutes the immune function. Thus, the contribution of HIV-1 to B-cell oncogenesis remains enigmatic. HIV-1 induces oxidative stress and DNA damage in infected cells via multiple mechanisms, including viral Tat protein. We have detected elevated levels of reactive oxygen species (ROS) and DNA damage in B-cells of HIV-infected individuals. As Tat is present in blood of infected individuals and is able to transduce cells, we hypothesized that it could induce oxidative DNA damage in B-cells promoting genetic instability and malignant transformation. Indeed, incubation of B-cells isolated from healthy donors with purified Tat protein led to oxidative stress, a decrease in the glutathione (GSH) levels, DNA damage and appearance of chromosomal aberrations. The effects of Tat relied on its transcriptional activity and were mediated by NF-κB activation. Tat stimulated oxidative stress in B-cells mostly via mitochondrial ROS production which depended on the reverse electron flow in Complex I of respiratory chain. We propose that Tat-induced oxidative stress, DNA damage and chromosomal aberrations are novel oncogenic factors favoring B-cell lymphomas in HIV-1 infected individuals.

Protective effect of Rhus coriaria fruit extracts against hydrogen peroxide-induced oxidative stress in muscle progenitors and zebrafish embryos.

BACKGROUND AND PURPOSE: Oxidative stress is involved in normal and pathological functioning of skeletal muscle. Protection of myoblasts from oxidative stress may improve muscle contraction and delay aging. Here we studied the effect of R. coriaria sumac fruit extract on human myoblasts and zebrafish embryos in conditions of hydrogen peroxide-induced oxidative stress. STUDY DESIGN AND METHODS: Crude ethanolic 70% extract (CE) and its fractions was obtained from sumac fruits. The composition of sumac ethyl acetate EtOAc fraction was studied by 1H NMR. The viability of human myoblasts treated with CE and the EtOAc fraction was determined by trypan blue exclusion test. Oxidative stress, cell cycle and adhesion were analyzed by flow cytometry and microscopy. Gene expression was analyzed by qPCR. RESULTS: The EtOAc fraction (IC50 2.57 µg/mL) had the highest antioxidant activity and exhibited the best protective effect against hydrogen peroxide-induced oxidative stress. It also restored cell adhesion. This effect was mediated by superoxide dismutase 2 and catalase. Pre-treatment of zebrafish embryos with low concentrations of the EtOAc fraction protected them from hydrogen peroxide-induced death in vivo. 1H NMR analysis revealed the presence of gallic acid in this fraction. CONCLUSION: Rhus coriaria extracts inhibited or slowed down the progress of skeletal muscle atrophy by decreasing oxidative stress via superoxide dismutase 2 and catalase-dependent mechanisms.

Hemisynthesis, Antitumoral Effect, and Molecular Docking Studies of Ferutinin and Its Analogues.

The natural product ferutinin was shown to act as an agonist to estrogen receptor ERα and agonist/antagonist to ERß featuring a weak antiproliferative activity toward breast cancer cells. To enhance this activity, ferutinin analogues were synthesized by esterification of jaeschkenadiol with different acids. These compounds were assayed for their in vitro antiproliferative activity against estrogen-dependent (MCF-7) and estrogen-independent (MDA-MB-231) breast cancer cell lines. Among the compounds, 3c' exhibited a potent inhibitory selective activity against MCF-7 with IC50 value of 1 µM. Docking simulation of 3c' in the ligand binding domain of the ERs indicated a potential antagonism interaction with both ER subtypes. Functional assay showed that 3c' binds as an antagonist to ERα protein while ferutinin acts as an agonist.

Berberis libanotica Ehrenb extract shows anti-neoplastic effects on prostate cancer stem/progenitor cells.

Cancer stem cells (CSCs), including those of advanced prostate cancer, are a suggested reason for tumor resistance toward conventional tumor therapy. Therefore, new therapeutic agents are urgently needed for targeting CSCs. Despite the minimal understanding of their modes of action, natural products and herbal therapies have been commonly used in the prevention and treatment of many cancers. Berberis libanotica Ehrenb (BLE) is a plant rich in alkaloids which may possess anti-cancer activity and a high potential for eliminating CSCs. We tested the effect of BLE on prostate cancer cells and our data indicated that this extract induced significant reduction in cell viability and inhibited the proliferation of human prostate cancer cell lines (DU145, PC3 and 22Rv1) in a dose- and time-dependent manner. BLE extract induced a perturbation of the cell cycle, leading to a G0-G1 arrest. Furthermore, we noted 50% cell death, characterized by the production of high levels of reactive oxidative species (ROS). Inhibition of cellular migration and invasion was also achieved upon treatment with BLE extract, suggesting a role in inhibiting metastasis. Interestingly, BLE extract had a major effect on CSCs. Cells were grown in a 3D sphere-formation assay to enrich for a population of cancer stem/progenitor cells. Our results showed a significant reduction in sphere formation ability. Three rounds of treatment with BLE extract were sufficient to eradicate the self-renewal ability of highly resistant CSCs. In conclusion, our results suggest a high therapeutic potential of BLE extract in targeting prostate cancer and its CSCs.

First computational evidence of a competitive stepwise and concerted mechanism for the reduction of antimalarial endoperoxides.

We study structural analogues of endoperoxides belonging to the family of G factors which present moderate to good antimalarial activity. Their biological activity is related to the reduction and cleavage of the O-O bond. Generally, the O-O bond reduction of model endoperoxides, as well as artemisinin, occurs by a concerted dissociative electron transfer (ET) mechanism. For the G3 and G3Me compounds, the experimental counterpart indicates an unexpected competition between a concerted and a stepwise mechanism, but no intermediate species can be isolated. We thus perform DFT studies on the reduction of G3 and G3Me compounds. We confirm the formation of an intermediate radical anion followed by cleavage of the O-O bond in a second step. We characterize the stable conformations for the radical anions G(3)(*-) and G(3)Me(*-) resulting from the ET and the associated reaction pathway. We also calculate the reorganization energy upon ET in relation to the Marcus theory using the DFT method. These results provide valuable insight into understanding the biological activity of G-factor endoperoxides as potential therapeutic antimalarial agents.

Electrochemical reduction of G3-factor endoperoxide and its methyl ether: evidence for a competition between concerted and stepwise dissociative electron transfer.

The reduction of the bicyclic G-factor endoperoxides G3 and G3Me was studied in N,N-dimethylformamide using cyclic voltammetry and convolution analysis. Electron transfer leads to irreversible cleavage of the O--O bond. Detailed analysis of the voltammetry curves reveals a non-linear dependence on the transfer coefficient indicating a mechanistic transition from a stepwise mechanism to one with more concerted character with increasing potential. By using quantum calculations to estimate the O--O bond dissociation energies, the experimental data was used to evaluate the standard reduction potentials and other pertinent thermochemical information.

Fe(II)-induced reduction of labelled endoperoxides. NMR degradation studies on G3 factor and its methyl ether.

The behavior of G3 factor and of its methylated or fluorinated analogues G3Me and G3F, was studied under Fe(II) conditions. Degradation products were isolated and characterized in each case. The use of labelled compounds allowed us to propose mechanisms in which a tertiary radical is involved. This radical rearranges by 5-exo-trig cyclization, or disproportionates in the case of G3Me. A correlation between antiplasmodial activity and stability of this radical is proposed.

Alkylation of natural endoperoxide G3-factor. Synthesis and antimalarial activity studies.

Alkylation of the peroxyhemiketal function is described and all synthesised endoperoxides show good antimalarial activity. New rearrangement reactions in the presence of CsCO3, and preliminary results on Fe(II) chemical reduction of the O-O bond are presented.

Crucial role of the peroxyketal function for antimalarial activity in the G-factor series.

New endoperoxides, related to the natural phytohormones known as G factors (G1, G2, G3), were modified on the side chain and the ketalic position. An unexpected rearrangement, specific to one diastereoisomer was observed in the deprotection step of O-silylated compounds and attributed to a hexacoordinated fluorosilicon intermediate. The reduction potential of these new peroxides was determined. They exhibited good to moderate antimalarial activity, greatly related to the presence of peroxyketal function.