Advanced bioengineering of male germ stem cells to preserve fertility.

In modern life, several factors such as genetics, exposure to toxins, and aging have resulted in significant levels of male infertility, estimated to be approximately 18% worldwide. In response, substantial progress has been made to improve in vitro fertilization treatments (e.g. microsurgical testicular sperm extraction (m-TESE), intra-cytoplasmic sperm injection (ICSI), and round spermatid injection (ROSI)). Mimicking the structure of testicular natural extracellular matrices (ECM) outside of the body is one clear route toward complete in vitro spermatogenesis and male fertility preservation. Here, a new wave of technological innovations is underway applying regenerative medicine strategies to cell-tissue culture on natural or synthetic scaffolds supplemented with bioactive factors. The emergence of advanced bioengineered systems suggests new hope for male fertility preservation through development of functional male germ cells. To date, few studies aimed at in vitro spermatogenesis have resulted in relevant numbers of mature gametes. However, a substantial body of knowledge on conditions that are required to maintain and mature male germ cells in vitro is now in place. This review focuses on advanced bioengineering methods such as microfluidic systems, bio-fabricated scaffolds, and 3D organ culture applied to the germline for fertility preservation through in vitro spermatogenesis.

Microfluidics-Integrated Sensors toward Rapid Detection of Single Nucleotide Variations.

Rapid detection of single nucleotide variations (SNVs) is of critical importance to early diagnosis of several diseases and the prediction of diverse responses to a specific treatment. Based on the information published in the literature, discrimination of SNVs is a developing area of study with great research enthusiasm and is also an area that can benefit from microfluidics-integrated designs. This review provides a brief overview of different microfluidics-based strategies for rapid detection of SNVs and mismatched bases. Sensors based on various microfluidic formats, such as paper-based microfluidic biosensors, droplet-based microfluidic systems, and magnetic bead-based microfluidic biosensors, have been discussed with respect to their specific pros and cons for SNV detection. These systems have shown promise for distributed on-site diagnostics in personalized medicine.

Nanophytomedicines for the Prevention of Metabolic Syndrome: A Pharmacological and Biopharmaceutical Review.

Metabolic syndrome includes a series of metabolic abnormalities that leads to diabetes mellitus and cardiovascular diseases. Plant extracts, due to their unique advantages like anti-inflammatory, antioxidant, and insulin sensitizing properties, are interesting therapeutic options to manage MetS; however, the poor solubility and low bioavailability of lipophilic bioactive components in the herbal extracts are two critical challenges. Nano-scale delivery systems are suitable to improve delivery of herbal extracts. This review, for the first time, focuses on nanoformulations of herbal extracts in MetS and related complications. Included studies showed that several forms of nano drug delivery systems such as nanoemulsions, solid lipid nanoparticles, nanobiocomposites, and green-synthesized silver, gold, and zinc oxide nanoparticles have been developed using herbal extracts. It was shown that the method of preparation and related parameters such as temperature and type of polymer are important factors affecting physicochemical stability and therapeutic activity of the final product. Many of these formulations could successfully decrease the lipid profile, inflammation, oxidative damage, and insulin resistance in in vitro and in vivo models of MetS-related complications. Further studies are still needed to confirm the safety and efficacy of these novel herbal formulations for clinical application.

Evaluation of the cytotoxic and apoptogenic effects of cinnamaldehyde on U87MG cells alone and in combination with doxorubicin.

BACKGROUND AND PURPOSE: In the present study, we tried for the first time to examine whether cinnamaldehyde (CA), with herbal nature, can be co-administrated with doxorubicin (DOX, as an anticancer drug) toward U87MG glioblastoma cells to potentiate its cytotoxic effect and overcome or reduce its side effects. EXPERIMENTAL APPROACH: The cytotoxic effect of DOX and CA, either individually or in combination, were evaluated on U87MG cells using the MTT method. The mechanism of action was studied by investigating the mode of cell death using caspase-3 and 9 activations, mitochondrial membrane potential (MMP) as well as sub G1 analysis. The expression of apoptosis- related genes (Bcl-2 and Bax) was also examined. FINDINGS / RESULTS: Cellular toxicity assay revealed that CA and DOX can potentially reduce the viability of U87MG cells with IC50 at 11.6 and 5 µg/mL, respectively. Exposure with the combination of CA and DOX significantly increased cytotoxic effect of DOX on U87MG cells. The results of SUBG1, MMP, and also caspase-3 and -9 activity assays, in association with the results corresponding to the Bax and Bcl-2 gene expressions, altogether revealed that CA can induce apoptosis on U87MG cells. Moreover, apoptogenic effects of DOX were found to be potentiated by CA. CONCLUSION AND IMPLICATIONS: The results of this study revealed the promising cytotoxic and apoptogenic role of CA on U87MG cells. Additionally, our findings demonstrated that CA is able to enhance the apoptosis induced by DOX on human glioblastoma cells. Collectively, these data suggested that co-exposure of CA and DOX could be effective for treatment of glioblastoma, but further in vivo and clinical studies are still needed to prove these results.

Microfluidic-assisted preparation of PLGA nanoparticles for drug delivery purposes: experimental study and computational fluid dynamic simulation.

This study, for the first time, tries to provide a simultaneous experimental and computational fluid dynamic (CFD) simulation investigation for production of uniform, reproducible, and stable polylactic-co-glycolic acid (PLGA) nanoparticles. CFD simulation was carried out to observe fluid flow behavior and micromixing in microfluidic system and improve our understanding about the governing fluid profile. The major objective of such effort was to provide a carrier for controlled and sustained release profile of different drugs. Different experimental parameters were optimized to obtain PLGA nanoparticles with proper size and minimized polydispersity index. The particle size, polydispersity, morphology, and stability of nanoparticles were compared. Microfluidic system provided a platform to control over the characteristics of nanoparticles. Using microfluidic system, the obtained particles were more uniform and harmonious in size, more stable, monodisperse and spherical, while particles produced by batch method were non-spherical and polydisperse. The best size and polydispersity index in the microfluidic method was obtained using 2% PLGA and 0.0625% (w/v) polyvinyl alcohol (PVA) solutions, and the flow rate ratio of 10:0.6 for PVA and PLGA solutions. CFD simulation demonstrated the high mixing intensity of about 0.99 at optimum condition in the microfluidic system, which is the possible reason for advantageous performance of this system. Altogether, the results of microfluidic-assisted method were found to be more reproducible, predictable, and controllable than batch method for producing a nanoformulation for delivery of drugs.

Evaluation of the Cytotoxic and Apoptogenic Effects of Glabridin and Its Effect on Cytotoxicity and Apoptosis Induced by Doxorubicin Toward Cancerous Cells.

Purposes: In the present study, we tried for the first time to examine the anti-proliferative and anti-apoptogenic effect of Glabridin (Glab) toward three groups of cancer cells (SKNMC, H1299, and A2780). Furthermore, the possibility of co-administration of Glab with doxorubicin (DOX) to these cells was also examined to find out whether Glab can potentiate the cytotoxic effect of this chemotherapy agent. Methods: Different cellular assays (MTT, caspase-3 activity, MMP, RT-PCR analysis) were carried out on the cancer cells treated with Glab. Results: Cellular toxicity assay revealed that Glab can potentially reduce the viability of these cells with IC50 concentrations up to 10, 12, and 38 µM toward A2780, SKNMC, and H1299 cell lines, respectively. The results of MMP and caspase-3 activity assays, in association with the results corresponding to the BAX and Bcl-2 gene expressions, altogether revealed that Glab can exert apoptogenic effect on these cells. The intrinsic mitochondrial pathway was found to be the main mechanism, in which Glab induced apoptosis toward H1299 cells and SKNMC cells, while the apoptosis mechanism for A2780 cells could be probably through extrinsic pathway. Glab also potentiated the cytotoxic effect of DOX and its accumulation in H1299 cell line. Conclusion: The results of this study revealed the promising cytotoxic role of Glab on different carcinoma cells. These data also suggested that co-chemotherapy method using Glab could be effective for treatment of cancer, but further in-vivo and clinical studies are still needed to assure these results.

Nanoformulations of natural products for management of metabolic syndrome.

Metabolic syndrome is a common metabolic disorder which has become a public health challenge worldwide. There has been growing interest in medications including natural products as complementary or alternative choices for common chemical therapeutics regarding their limited side effects and ease of access. Nanosizing these compounds may help to increase their solubility, bioavailability, and promisingly enhance their efficacy. This study, for the first time, provides a comprehensive overview of the application of natural-products-based nanoformulations in the management of metabolic syndrome. Different phytochemicals including curcumin, berberine, Capsicum oleoresin, naringenin, emodin, gymnemic acid, resveratrol, quercetin, scutellarin, stevioside, silybin, baicalin, and others have been nanosized hitherto, and their nanosizing method and effect in treatment and alleviating metabolic syndrome have been reviewed and discussed in this study. It has been discovered that there are several pathways or molecular targets relevant to metabolic disorders which are affected by these compounds. Various natural-based nanoformulations have shown promising effect in treatment of metabolic syndrome, and therefore can be considered as future candidates instead of or in conjunction with pharmaceutical drugs if they pass clinical trials successfully.

Hesperidin as a Neuroprotective Agent: A Review of Animal and Clinical Evidence.

Neuroprotection is the preservation of function and networks of neural tissues from damages caused by various agents, as well as neurodegenerative diseases such as Parkinson's, Alzheimer's, Huntington's diseases, and multiple sclerosis. Hesperidin, a flavanone glycoside, is a natural phenolic compound with a wide range of biological effects. Mounting evidence has demonstrated that hesperidin possesses inhibitory effect against development of neurodegenerative diseases. Our review discusses neuropharmacological mechanisms for preventive and therapeutic effects of hesperidin in neurodegenerative diseases. In addition, the review examines clinical evidence confirming its neuroprotective function. Various cellular and animal models specific to neurodegenerative diseases have been conducted to evaluate the underlying neuropharmacological mechanisms of hesperidin. Neuroprotective potential of this flavonoid is mediated by improvement of neural growth factors and endogenous antioxidant defense functions, diminishing neuro-inflammatory and apoptotic pathways. Despite the various preclinical studies on the role of hesperidin in the neurodegenerative diseases, less is known about its definite effect on humans. A limited number of clinical trials showed that hesperidin-enriched dietary supplements can significantly improve cerebral blood flow, cognition, and memory performance. Further clinical trials are also required for confirming neuroprotective efficacy of this natural flavonoid and evaluating its safety profile.

Acute and Subchronic Toxicological Evaluations of Allium rotundum L.: A Dietary Plant from Iran.

Allium rotundum L. is a dietary plant with diverse nutritional and herbal applications. According to its widespread application in Iranians' diets, understanding the possible adverse effects and toxic activities could be of major importance. The aim of this study was to establish the acute and subchronic toxicity profile of the hydroalcoholic extract of Allium rotundum on male and female Wistar rats. The acute study indicated no adverse effect or toxic activity after administration of the extract, suggesting that the LD50 value is up to 5,000 mg/kg body weight for the extract. The subchronic study at three doses (250, 500, and 750 mg/kg body weight/day) supported the results of acute study and revealed that no abnormal change or toxicity was induced by the extract in both male and female Wistar rats. All the biochemical and hematological parameters of the treated rats were in historical range after long-term administration of the extract. The histopathological examination also revealed no lesion or alteration in the tissue of vital organs (kidney, liver, heart, lung, and spleen). The NOAEL (no observed adverse effect level) value was high enough (greater than 750 mg/kg body weight/ day) to conclude the nontoxic nature of this extract. The safety of this extract was affirmed by the acute and subchronic toxic studies and suggested that this plant could be a proper and effective dietary plant due to its high nutritive value and inherent therapeutic properties.

Natural product-based nanomedicines for wound healing purposes: therapeutic targets and drug delivery systems.

Wound healing process is an intricate sequence of well-orchestrated biochemical and cellular phenomena to restore the integrity of the skin and subcutaneous tissue. Several plant extracts and their phytoconstituents are known as a promising alternative for wound healing agents due to the presence of diverse active components, ease of access, and their limited side effects. The development of nanotechnological methods can help to improve the efficacy of different therapeutics as well as herbal-based products. Here, we present a review of the efficacy of the plant based-nanomaterials in the management of wounds and discuss the involved therapeutic targets. For this purpose, a profound search has been conducted on in vitro, in vivo, and/or clinical evidences evaluating the efficacy and pharmacological mechanisms of natural product-based nanostructures on different types of wounds. Different pharmacological targets are involved in the wound healing effects of herbal-based nanostructures, including suppressing the production of inflammatory cytokines and inflammatory transduction cascades, reducing oxidative factors and enhancing antioxidative enzymes, and promoting neovascularization and angiogenic pathways through increasing the expression of vascular endothelial growth factor, fibroblast growth factor, and platelet-derived growth factor. Moreover, nanostructure of plant extracts and their phytochemicals can enhance their bioavailability, control their release in the form of sustained delivery systems to the wound site, and enhance the permeability of these therapeutics to the underlying skin layers, which are all necessary for the healing process. Overall, various plant extracts and their natural compounds, used in nanoformulations, have demonstrated high activity in the management of wounds and thus can be assumed as future pharmaceutical drugs.

Green Synthesis of Carbon Dots Derived from Walnut Oil and an Investigation of Their Cytotoxic and Apoptogenic Activities toward Cancer Cells.

Purpose: This paper introduces a green and simple hydrothermal synthesis to prepare carbon quantum dots (CQDs) from walnut oil with a high quantum yield. In addition, cytotoxic and apoptogenic properties of the CQDs were analyzed on human cancer cell lines. Methods: The optical properties and morphological characteristic were investigated by the TEM, XRD, FT-IR, UV-vis and photoluminescence (PL).The cytotoxic potential of walnut CQDs was evaluated on PC3, MCF-7 and HT-29 human carcinoma cell lines using the MTT methods. The mechanism of action was studied by investigating the mode of cell death using the activation of caspase-3 and 9 as well as mitochondrial membrane potential (MMP). Cellular uptake of the CQDs was detected by fluorescence microscope. CQDs had an average size of 12 nm and a significant emission at 420 nm at an excitation wavelength of 350 nm was recorded. Results: The prepared CQDs possessed a good fluorescent quantum yield of 14.5% with quinine sulfate (quantum yield 54%) as a reference and excellent photo as well as pH stabilities. The walnut CQDs were proved to be an extremely potent cytotoxic agent, especially against MCF-7 and PC-3 cell lines. Induction of apoptosis by CQDs was accompanied by an increase in the activation of caspase-3. Caspase-9 activity did not increase after exposure to the CQDs. Additionally; the MMP did not show any significant loss. Conclusion: The results of our study can corroborate the cytotoxic and apoptotic effect of walnut CQDs in the PC3 and MCF-7 cancer cell lines.

Dietary Plants for the Prevention and Management of Kidney Stones: Preclinical and Clinical Evidence and Molecular Mechanisms.

Kidney stones are one of the oldest known and common diseases in the urinary tract system. Various human studies have suggested that diets with a higher intake of vegetables and fruits play a role in the prevention of kidney stones. In this review, we have provided an overview of these dietary plants, their main chemical constituents, and their possible mechanisms of action. Camellia sinensis (green tea), Rubus idaeus (raspberry), Rubia cordifolia (common madder), Petroselinum crispum (parsley), Punica granatum (pomegranate), Pistacia lentiscus (mastic), Solanum xanthocarpum (yellow-fruit nightshade), Urtica dioica (stinging nettle), Dolichos biflorus (horse gram), Ammi visnaga (khella), Nigella sativa (black-cumin), Hibiscus sabdariffa (roselle), and Origanum vulgare (oregano) have received considerable interest based on scientific evidence. Beside these dietary plants, phytochemicals-such as catechin, epicatechin, epigallocatechin-3-gallate, diosmin, rutin, quercetin, hyperoside, and curcumin-as antioxidant dietary phyto-phenols were found to be effective for the prevention of urolithiasis (the process of stone formation in the urinary tract). The main underlying mechanisms of these dietary plants and their isolated phytonutrients in the management of urolithiasis include diuretic, antispasmodic, and antioxidant activity, as well as an inhibitory effect on crystallization, nucleation, and aggregation of crystals. The results as presented in this review demonstrate the promising role of dietary plants and phytophenols in the prevention and management of kidney stones. Further investigations are required to confirm the safety and efficacy of these compounds.

The Protective Effect of Different Extracts of Three Artemisia Species against H2O2-Induced Oxidative Stress and Apoptosis in PC12 Neuronal Cells.

BACKGROUND: Oxidative stress causes cell damage and is involved in many neurological diseases. The antioxidant properties of plant materials for the maintenance of health and protecting against different diseases stimulated scientist to investigate different herbs. Different Artemisia species have exhibited antioxidant activity. This study aims to investigate whether different Artemisia species could protect the PC12 cells against oxidative stress mediated by H2O2. METHODS: For this purpose, different extracts of three Artemisia species (Artemisia aucheri, Artemisia turanica, and Artemisia turcomanica) were prepared using petroleum ether, dichloromethane, ethyl acetate, ethanol, and Water: Ethanol mixture (1:1 volume ratio). The protective effect of the prepared extracts against H2O2-induced cytotoxicity and reactive oxygen species production were compared. The effect of treatment of PC12 cells with different extracts on total glutathione (GSH) level, caspase-3 activity, and mitochondrial membrane potential were also compared. RESULTS: The A. aucheri extracts could not rescue the PC12 cells from oxidative stress consequences. The A. turanica and A. turcomanica extracts were found potent in suppressing the toxicity and apoptosis of PC12 cells mediated by H2O2 and significantly antagonized the H2O2-induced GSH depletion. The hydroethanolic and ethyl acetate extracts of A. turanica and the petroleum ether and hydroethanolic extracts of A. turcomanica more efficiently suppressed cytotoxicity and loss of GSH caused by H2O2. CONCLUSION: This study shows the protective effects of Artemisia extracts on PC12 cell line and suggested that these species could be also considered as promising neuroprotective agents in treatment of different neurodegenerative diseases. SUMMARY: Artemisia turanica and Artemisia turcomanica extracts were found to potentially exert neuroprotective effect on PC12 cells. The results exhibited that the cytoprotective potential and anti-apoptotic mechanism of these species is not the same for different extracts, and suggested that based on the type of species and the type of solvents used in extraction, both intrinsic and extrinsic pathways could be included in the anti-apoptotic mechanism of these species. Abbreviations Used: GSH: Glutathion. ROS: Reactive Oxygen Species. GSSG: Glutathione disulfide. DCF-DA:2',7'-Dichlorofluorescin diacetate. FBS: Fetal Bovin Serum. MMP: Mitochondrial Membrane Potential. H-Et: Hydro-ethanolic. DCM: Dichloromethane. PE: Petroleum Ether. Et: Etanolic. EA: Ethyl Acetate.

Preparation and characterization of silk fibroin hydrogel as injectable implants for sustained release of Risperidone.

The principal objective of the present study is to achieve a depot formulation of Risperidone by gelation of silk fibroin (SF). For this purpose, hydrochloric acid (HCl)/acetone-based and methanol-based hydrogels were prepared with different drug/polymer ratios (1:3, 1:6, and 1:15). For all the drug-loaded methanol-based hydrogels, gel transition of SF solutions occurred immediately and the gelation time was 1 min, while the gelation time of HCL/acetone-based hydrogels was around 360 min. According to the results obtined from Fourier transform infrared spectroscopy (FT-IR) and X-ray diffraction (XRD) spectra, solvent systems and Risperidone could induce ß-sheet structure, but HCL/acetone system had the lowest effect on induction of ß-sheets. The crystallinity was increased by increasing the amount of Risperidone, and drug to polymer ratio of 1:3 possessed the highest crystallinity. Thermogravimetric analysis (TGA) indicated that increasing the amount of drug in formulation increased the stability of hydrogels, and methanol-based hydrogel with a ratio of 1:3 had the most stable structure. The release rate of Risperidone from methanol-based hydrogel at ratio of 1:3 was lower than that for HCl/acetone-based one, and it decreased by increasing the amount of Risperidone. The release of Risperidone from methanol hydrogel at ratios 1:3 and 1:6 continued up to 25 d which is acceptable for depot form of Risperidone and shows that the extended release of Risperidone was achieved successfully. In conclusion, SF hydrogel with the ability to respond to the environmental stimuli is an excellent candidate for injectable implants for extended release of Risperidone.

Preparation and in vitro characterization of retinoic acid-loaded poly(ε-caprolactone)-poly(ethylene glycol)-poly(ε-caprolactone) micelles.

In order to achieve the controlled release of all-trans-retinoic acid (ATRA), poly(ε-caprolactone)-poly(ethylene glycol)-poly(ε-caprolactone) (PCL-PEG-PCL) copolymer with average molecular weight of 5.34 kDa was synthesized. The nanosized micelles were prepared from copolymer by nano-precipitation method. Critical association concentration (CAC) of micelles was measured by fluorimetry and results indicated low CAC value of micelles (1.9 × 10-3 g/L). ATRA was encapsulated in the core of micelles using different ratios of drug to copolymer. In the case of 10% drug to polymer ratio, more than 80% of the drug was released within 3 days, whereas for ratio of 2% more than 90% of the drug was released within 3 h. The cytotoxic study performed by MTT assay showed that H1299 survival percent decreased significantly (P ≤ 0.05) after exposure to drug-loaded micelles, while no proliferation inhibition effect was observed by either free ATRA or blank PCL-PEG-PCL micelles.

Preparation and Characterization of Pistacia khinjuk Gum Nanoparticles Using Response Surface Method: Evaluation of Its Anti-Bacterial Performance and Cytotoxicity.

Purpose: This study aims to prepare a novel, natural nanoparticle (NP) as a drug carrier, which also has inherent therapeutic effects. Methods: Pistacia khinjuk gum NPs were prepared and Response surface methodology (RSM) was used for statistical analysis of data and optimizing the size of NPs. Results: NPs were in the range of 75.85-241.3 nm. The optimization study was carried out, and an optimized size (70.86nm) was obtained using DMSO as a solvent. The volume of the organic phase was 111.25µl, and the concentration of gum was 1% w/v. The cell viability assay was performed on the pure gum and NPs toward ß-TC3, MCF7, and HT29 cell lines. It was observed that NPs have higher cytotoxic activity in comparison with pure gum, and that the IC50value was achieved at 1% of NPs in ß-TC3 cells. The obtained NPs demonstrated antibacterial activity against two bacterial strains (Pseudomonas aeruginosa and Staphylococcus aureus). Conclusion: Altogether, according to the obtained results, these NPs with inherent cytotoxicity and antibacterial activity are an attractive carrier for drug delivery.

Preparation of a reproducible long-acting formulation of risperidone-loaded PLGA microspheres using microfluidic method.

The aim of the present study is to prepare risperidone-loaded poly lactic-co-glycolic acid (PLGA) microspheres within microfluidic system and to achieve a formulation with uniform size and monotonic and reproducible release profile. In comparison to batch method, T-junction and serpentine chips were utilized and optimizing study was carried out at different processing parameters (e.g. PLGA and surfactant concentration and flow rates ratio of outer to inner phase). The computational fluid dynamic (CFD) modeling was performed, and loading and release study were carried out. CFD simulation indicates that increasing the flow rate of aqueous phase cause to decrease the droplet size, while the change in size of microspheres did not follow a specific pattern in the experimental results. The most uniform microspheres and narrowest standard deviation (66.79 µm ± 3.32) were achieved using T-junction chip, 1% polyvinylalcohol, 1% PLGA and flow rates ratio of 20. The microfluidic-assisted microspheres were more uniform with narrower size distribution. The release of risperidone from microspheres produced by the microfluidic method was more reproducible and closer to zero-order kinetic model. The release profile of formulation with 2:1 drug-to-polymer ratio was the most favorable release, in which 41.85% release could be achieved during 24 days.