Tissue clearing and three-dimensional imaging of intact tissues: a review on FACT protocol.

FACT is a developed technique for clearing tissues that does not use acrylamide. Since the removal of lipids is crucial for transparency and efficient antibody staining throughout the tissue, especially for microscopy and imaging, it is a harmful process that can cause the loss of important biological molecules such as proteins. The FACT technique overcomes this by chemically bonding the membrane and intracellular proteins with the extracellular matrix, creating a massive 3D hydrogel matrix and providing structural support to fortify the tissue during processing. Compared to other acrylamide-based techniques, the FACT technique requires less labor and harmful chemicals and is therefore considered a more suitable option. In this study, we describe the complete FACT protocol for antibody staining and imaging of whole-cleared tissues while preserving structure and improving image quality. The protocol includes tissue perfusion, fixation, clearing, antibody staining, refractive index matching (RIM) (), microscopy, and imaging. The timing for each step varies depending on the size, weight, and type of tissue, as well as the type of immunostaining. We provide an example of the FACT protocol using mouse brain tissue, which demonstrates its suitability for molecular interrogation analysis of large tissues. The FACT technique has been successfully performed on different types of tissues, making it a favorable choice for a variety of applications.

Comprehensive Overview of Innovative Strategies in Preventing Renal Ischemia-reperfusion Injury: Insights from Bibliometric and In Silico Analyses.

BACKGROUND: Ischemia-Reperfusion Injury (IRI) is a complex pathophysiological process with severe consequences, including irreversible loss of renal function. Various intraoperative prevention methods have been proposed to mitigate the harmful effects of warm ischemia and kidney reperfusion. AIM: This comprehensive analysis provides an overview of pharmacological agents and intraoperative methods for preventing and treating renal IRI. METHODS: Our analysis revealed that eplerenone exhibited the highest binding affinity to crucial targets, including Aldehyde Dehydrogenase (AD), Estrogen Receptor (ER), Klotho protein, Mineralocorticoid Receptor (MR), and Toll-Like Receptor 4 (TLR4). This finding indicates eplerenone's potential as a potent preventive agent against IRI, surpassing other available therapeutics like Benzodioxole, Hydrocortisone, Indoles, Nicotinamide adenine dinucleotide, and Niacinamide. In preventing kidney IRI, our comprehensive analysis emphasizes the significance of eplerenone due to its strong binding affinity to key targets involved in the pathogenesis of IRI. RESULTS: This finding positions eplerenone as a promising candidate for further clinical investigation and consideration for future clinical practice. CONCLUSION: The insights provided in this analysis will assist clinicians and researchers in selecting effective preventive approaches for renal IRI in surgical settings, potentially improving patient outcomes.

Prokaryotic microvesicles Ortholog of eukaryotic extracellular vesicles in biomedical fields.

Every single cell can communicate with other cells in a paracrine manner via the production of nano-sized extracellular vesicles. This phenomenon is conserved between prokaryotic and eukaryotic cells. In eukaryotic cells, exosomes (Exos) are the main inter-cellular bioshuttles with the potential to carry different signaling molecules. Likewise, bacteria can produce and release Exo-like particles, namely microvesicles (MVs) into the extracellular matrix. Bacterial MVs function with diverse biological properties and are at the center of attention due to their inherent therapeutic properties. Here, in this review article, the comparable biological properties between the eukaryotic Exos and bacterial MVs were highlighted in terms of biomedical application. Video Abstract.

Exosomes as Theranostic Agents in Reproduction System.

Exosomes (Exos), belonging to extracellular vesicles, are cell-derived nano-sized vesicles with the potential to carry different kinds of biological molecules. Many studies have proved the impacts of exosomal cargo on several biological processes in female and male reproductive systems. It is also hypothesized that changes in exosomal cargo are integral to the promotion of certain pathological conditions, thus Exos can be used as valid biomarkers for the diagnosis of infertility and other abnormal conditions. Here, efforts are made to collect some recent data related to the physiological significance of Exos in the reproductive system, and their potential therapeutic effects. It is anticipated that the current review article will lay the groundwork for elucidating the source and mechanisms by which Exos control the reproductive system additionally supplying fresh methods and concepts for the detection and treatment of disorders associated with fertility for future studies.

Comparison the therapeutic effects of bone marrow CD144+ endothelial cells and CD146+ mesenchymal stem cells in POF rats.

Introduction: Premature ovarian insufficiency (POI) is a challenging issue in terms of reproduction biology. In this study, therapeutic properties of bone marrow CD146+ mesenchymal stem cells (MSCs) and CD144+ endothelial cells (ECs) were separately investigated in rats with POI. Methods: POI rats were classified into control POI, POI + CD146+ MSCs, and POI + CD144+ ECs groups. Enriched CD146+ MSCs and CD144+ ECs were directly injected into ovarian tissue (15 × 104 cells/10 µL) in relevant groups. After 4 weeks, follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) levels were measured in blood samples. Ovarian tissues were collected and subjected to Hematoxylin-Eosin and Masson's trichrome staining. The expression of angp-2, vegfr-2, smad-2, -4, -6, and tgf-ß1 was studied using qRT-PCR analysis. Histopathological examination indicated an increased pattern of atretic follicles in the POI group related to the control rats (P<0.0001). Results: Data indicated that injection of POI + CD146+ MSCs and CD144+ ECs in POI rats reduced atretic follicles and increased the number of normal follicles (P<0.01). Along with these changes, the content of blue-colored collagen fibers was diminished after cell transplantation. Besides, cell transplantation in POI rats had the potential to reduce increased FSH, and LH levels (P<0.05). In contrast, E2 content was increased in POI + CD146+ MSCs and POI + CD144+ ECs groups compared to control POI rats, indicating restoration of follicular function. CD144+ (smad-2, and -4) and CD146+ (smad-6) cells altered the activity of genes belonging TGF-ß signaling pathway. Unlike POI + CD146+ MSCs, aberrant angiogenesis properties were significantly down-regulated in POI + CD144+ ECs related to the control POI group (P<0.05). Conclusion: The transplantation of bone marrow CD146+ and CD144+ cells can lead to the restoration of ovarian tissue function in POI rats via modulating different mechanisms associated with angiogenesis and fibrosis.

Synergistic effect of chitosan-alginate composite hydrogel enriched with ascorbic acid and alpha-tocopherol under hypoxic conditions on the behavior of mesenchymal stem cells for wound healing.

BACKGROUND: In regenerative medicine, especially skin tissue engineering, the focus is on enhancing the quality of wound healing. Also, several constructs with different regeneration potentials have been used for skin tissue engineering. In this study, the regenerative properties of chitosan-alginate composite hydrogels in skin wound healing under normoxic and hypoxic conditions were investigated in vitro. METHODS: The ionic gelation method was used to prepare chitosan/alginate (CA) hydrogel containing CA microparticles and bioactive agents [ascorbic acid (AA) and α-tocopherol (TP)]. After preparing composite hydrogels loaded with AA and TP, the physicochemical properties such as porosity, pore size, swelling, weight loss, wettability, drug release, and functional groups were analyzed. Also, the hemo-biocompatibility of composite hydrogels was evaluated by a hemolysis test. Then, the rat bone marrow mesenchymal stem cells (rMSCs) were seeded onto the hydrogels after characterization by flow cytometry. The survival rate was analyzed using MTT assay test. The hydrogels were also investigated by DAPI and H&E staining to monitor cell proliferation and viability. To induce hypoxia, the cells were exposed to CoCl2. To evaluate the regenerative potential of rMSCs cultured on CA/AA/TP hydrogels under hypoxic conditions, the expression of the main genes involved in the healing of skin wounds, including HIF-1α, VEGF-A, and TGF-ß1, was investigated by real-time PCR. RESULTS: The results demonstrated that the prepared composite hydrogels were highly porous, with interconnected pores that ranged in sizes from 20 to 188 µm. The evaluation of weight loss showed that the prepared hydrogels have the ability to biodegrade according to the goals of wound healing. The reduction percentage of CA/AA/TP mass in 21 days was reported as 21.09 ± 0.52%. Also, based on wettability and hemolysis tests of the CA/AA/TP, hydrophilicity (θ = 55.6° and 53.7°) and hemocompatibility with a hemolysis ratio of 1.36 ± 0.19 were evident for them. Besides, MTT assay, DAPI, and H&E staining also showed that the prepared hydrogels provide a suitable substrate for cell growth and proliferation. Finally, based on real-time PCR, increased expression levels of VEGF and TGF-ß1 were observed in rMSCs in hypoxic conditions cultured on the prepared hydrogels. CONCLUSIONS: In conclusion, this study provides evidence that 3D CA/AA/TP composite hydrogels seeded by rMSCs in hypoxic conditions have great potential to improve wound healing.

Phenolated alginate hydrogel induced osteogenic properties of mesenchymal stem cells via Wnt signaling pathway.

Osteogenic properties of phenolated alginate (1.2 %) hydrogel containing collagen (0.5 %)/nano-hydroxyapatite (1 %) were studied on human mesenchymal stem cells in vitro. The phenolation rate and physical properties of the hydrogel were assessed using nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FTIR), Scanning electron microscope (SEM), swelling ratio, gelation time, mechanical assay, and degradation rate. The viability of encapsulated cells was monitored on days 7, 14, and 21 using an MTT assay. Osteoblast differentiation was studied using western blotting, and real-time PCR. Using PCR array analysis, the role of the Wnt signaling pathway was also investigated. Data showed that the combination of alginate/collagen/nanohydroxyapatite yielded proper mechanical features. The addition of nanohydroxyapatite, and collagen reduced degradation, swelling rate coincided with increased stiffness. Elasticity and pore size were also diminished. NMR and FTIR revealed suitable incorporation of collagen and nanohydroxyapatite in the structure of alginate. Real-time PCR analysis and western blotting indicated the expression of osteoblast-related genes such as Runx2 and osteocalcin. PCR array revealed the induction of numerous genes related to Wnt signaling pathways during the maturation of human stem cells toward osteoblast-like cells. In vivo data indicated that transplantation of phenolated alginate/collagen/nanohydroxyapatite hydrogel led to enhanced de novo bone formation in rats with critical-sized calvarial defects. Phenolated alginate hydrogel can promote the osteogenic capacity of human amniotic membrane mesenchymal stem cells in the presence of nanohydroxyapatite and collagen via engaging the Wnt signaling pathway.

Enhancing differentiation of menstrual blood-derived stem cells into female germ cells using a bilayer amniotic membrane and nano-fibrous fibroin scaffold.

Three-dimensional nanofiber scaffolds offer a promising method for simulating in vivo conditions within the laboratory. This study aims to investigate the influence of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold on the differentiation of human menstrual blood mesenchymal stromal/stem cells (MenSCs) into female germ cells. MenSCs were isolated and assigned to four culture groups: (i) MenSCs co-cultured with granulosa cells (GCs) using the scaffold (3D-T group), (ii) MenSCs using the scaffold alone (3D-C group), (iii) MenSCs co-cultured only with GCs (2D-T group), and (iv) MenSCs without co-culture or scaffold (2D-C group). Both MenSCs and GCs were independently cultured for two weeks before co-culturing was initiated. Flow cytometry was employed to characterize MenSCs based on positive markers (CD73, CD90, and CD105) and negative markers (CD45 and CD133). Additionally, flow cytometry and immunocytochemistry were used to characterize the GCs. Differentiated MenSCs were analyzed using real-time PCR and immunostaining. The real-time PCR results demonstrated significantly higher levels of VASA expression in the 3D-T group compared to the 3D-C, 2D-T, and 2D-C groups. Similarly, the SCP3 mRNA level in the 3D-T group was notably elevated compared to the 3D-C, 2D-T, and 2D-C groups. Moreover, the expression of GDF9 was significantly higher in the 3D-T group when compared to the 3D-C, 2D-T, and 2D-C groups. Immunostaining results revealed a lack of signal for VASA, SCP3, or GDF9 markers in the 2D-T group, while some cells in the 3D-T group exhibited positive staining for all these proteins. These findings suggest that the combination of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold with co-culturing GCs facilitates the differentiation of MenSCs into female germ cells.

Microbial Landscape and Antibiotic-Susceptibility Profiles of Microorganisms in Patients with Bacterial Pneumonia: A Comparative Cross-Sectional Study of COVID-19 and Non-COVID-19 Cases in Aktobe, Kazakhstan.

This cross-sectional study investigated the microbial landscape and antibiotic-resistance patterns in patients with bacterial pneumonia, with a focus on the impact of COVID-19. Sputum samples from individuals with bacterial pneumonia, including coronavirus disease 2019-positive polymerase chain reaction (COVID-19-PCR+), COVID-19-PCR- and non-COVID-19 patients, were analyzed. Surprisingly, the classic etiological factor of bacterial pneumonia, Streptococcus pneumoniae, was rarely isolated from the sputum samples. Furthermore, the frequency of multidrug-resistant pathogens was found to be higher in non-COVID-19 patients, highlighting the potential impact of the pandemic on antimicrobial resistance. Strains obtained from COVID-19-PCR+ patients exhibited significant resistance to commonly used antibiotics, including fluoroquinolones and cephalosporins. Notably, the ESKAPE pathogens, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter cloacae, and Enterobacter aerogenes, were identified among the isolated microorganisms. Our findings underscore the urgent need for infection control measures and responsible antibiotic use in healthcare settings, as well as the importance of enhancing pneumonia diagnostics and implementing standardized laboratory protocols.

Dysregulated miRNAs in recurrent miscarriage: A systematic review.

Recurrent miscarriage (RM) is a complex reproductive medicine disease that affects many families. The cause of RM is unclear at this time; however, lifestyle and genetic variables may influence the process. The slight alteration in miRNA expression has enormous consequences for a variety of difficulties, one of which may be RM. The target of this systematic study was to provide a framework of the dysregulated miRNAs in RM. The Prisma guidelines were applied to perform current systematic review pertaining to articles in the seven databases. Thirty-nine papers out of 245 received fulfilled all inclusion requirements. From all the mentioned miRNAs, 40 were up-regulated (65.57 %), whereas 21 were down-regulated (34.43 %). These dysregulated miRNAs contributed to the pathophysiology of RM by influencing key pathways and processes such as apoptosis, angiogenesis, epithelial-mesenchymal transition, and the immune system. Understanding the dysregulation of miRNAs, as well as the pathways and processes that engage these miRNAs and impact disease pathogenesis, may aid in clarifying the unknown underlying mechanisms of RM and the development of novel molecular therapeutic targets and medical domains.

Interaction of lncRNAs with mTOR in colorectal cancer: a systematic review.

Colorectal cancer (CRC) is the third most widespread cancer and the fourth leading lethal disease among different societies. It is thought that CRC accounts for about 10% of all newly diagnosed cancer cases with high-rate mortality. lncRNAs, belonging to non-coding RNAs, are involved in varied cell bioactivities. Emerging data have confirmed a significant alteration in lncRNA transcription under anaplastic conditions. This systematic review aimed to assess the possible influence of abnormal mTOR-associated lncRNAs in the tumorigenesis of colorectal tissue. In this study, the PRISMA guideline was utilized based on the systematic investigation of published articles from seven databases. Of the 200 entries, 24 articles met inclusion criteria and were used for subsequent analyses. Of note, 23 lncRNAs were prioritized in association with the mTOR signaling pathway with up-regulation (79.16%) and down-regulation (20.84%) trends. Based on the obtained data, mTOR can be stimulated or inhibited during CRC by the alteration of several lncRNAs. Determining the dynamic activity of mTOR and relevant signaling pathways via lncRNAs can help us progress novel molecular therapeutics and medications.

Melatonin blunted the angiogenic activity in 3D colon cancer tumoroids by the reduction of endocan.

BACKGROUND: Complexity and heterogeneity of the tumor niche are closely associated with the failure of therapeutic protocols. Unfortunately, most data have been obtained from conventional 2D culture systems which are not completely comparable to in vivo microenvironments. Reconstructed 3D cultures composed of multiple cells are valid cell-based tumor models to recapitulate in vivo-like interaction between the cancer cells and stromal cells and the oncostatic properties of therapeutics. Here, we aimed to assess the tumoricidal properties of melatonin on close-to-real colon cancer tumoroids in in vitro conditions. METHODS: Using the hanging drop method, colon cancer tumoroids composed of three cell lines, including adenocarcinoma HT-29 cells, fibroblasts (HFFF2), and endothelial cells (HUVECs) at a ratio of 2: 1: 1, respectively were developed using 2.5% methylcellulose. Tumoroids were exposed to different concentrations of melatonin, from 0.005 to 0.8 mM and 4 to 10 mM, for 48 h. The survival rate was measured by MTT and LDH leakage assays. Protein levels of endocan and VEGF were assessed using western blotting. Using histological examination (H & E) staining, the integrity of cells within the tumoroid parenchyma was monitored. RESULTS: Despite the reduction of viability rate in lower doses, the structure of tumoroids remained unchanged. In contrast, treatment of tumoroids with higher doses of melatonin, 4 and 10 mM, led to disaggregation of cells and reduction of tumoroid diameter compared to the non-treated control tumoroids (p < 0.05). By increasing melatonin concentration from 4 to 10 mM, the number of necrotic cells increased. Data showed the significant suppression of endocan in melatonin-treated tumoroids related to the non-treated controls (p < 0.05). According to our data, melatonin in higher doses did not alter protein levels of VEGF (p > 0.05). CONCLUSIONS: Melatonin can exert its tumoricidal properties on colon cancer tumoroids via the reduction of tumor cell viability and inhibition of the specific pro-angiogenesis factor.

Amniotic fluid-derived exosomes attenuated fibrotic changes in POI rats through modulation of the TGF-ß/Smads signaling pathway.

In the current study, we investigated the regenerative effects of amniotic fluid exosomes (AF-Exos) in a rat model for premature ovarian insufficiency (POI). POI is a condition characterized by a decrease in ovarian function that can lead to infertility. We induced POI by administering cyclophosphamide (CTX) for 15 consecutive days, and then transplanted AF-Exos directly into both ovarian tissues. Four weeks later, we measured the serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2), and performed histopathological evaluations using H & E and Masson's trichrome staining. We also monitored the expression of genes related to the TGF-ß signaling pathway using real-time PCR and examined the fertility rate of POI rats after AF-Exos therapy. Histological analysis showed an increase in atretic follicles and a decrease in healthy follicle count after POI induction. Four weeks post-AF-Exos intervention, the healthy follicle count increased (p < 0.01) while the atretic follicle count decreased (p < 0.001). In parallel, the deposition of collagen fibers also decreased following AF-Exos transplantation. The concentrations of FSH and LH hormones in sera remained unchanged after injection of AF-Exos, while E2 levels increased (p < 0.05). The expression of Smad-4 (p < 0.01) and Smad-6 (p < 0.05) was upregulated in POI rats that received AF-Exos, while Smad-2, TGF-ß1, TNF-α, and IL-10 remained statistically unchanged. Our records showed a notable increase in litter number after AF-Exos compared to the non-treated POI rats. These results suggest that AF-Exos transplantation has the potential to restore ovarian function through the TGF-ß/Smads signaling pathway in POI rats.

A Bibliometric and In Silico-Based Analysis of Anti-Lung Cancer Compounds from Sea Cucumber.

Lung cancer is one of the most lethal malignancies in the world. However, current curative approaches for treating this type of cancer have some weaknesses. Therefore, scientists are attempting to discover new anti-lung cancer agents. Sea cucumber is a marine-derived source for discovering biologically active compounds with anti-lung cancer properties. To explore the anti-lung cancer properties of sea cucumber, we analyzed surveys using VOSviewer software and identified the most frequently used keywords. We then searched the Google Scholar database for compounds with anti-lung cancer properties within that keyword family. Finally, we used AutoDock 4 to identify the compounds with the highest affinity for apoptotic receptors in lung cancer cells. The results showed that triterpene glucosides were the most frequently identified compounds in studies examining the anti-cancer properties of sea cucumbers. Intercedenside C, Scabraside A, and Scabraside B were the three triterpene glycosides with the highest affinity for apoptotic receptors in lung cancer cells. To the best of our knowledge, this is the first time that anti-lung cancer properties of sea cucumber-derived compounds have been examined in in silico conditions. Ultimately, these three components displayed anti-lung cancer properties in in silico conditions and may be used for the manufacture of anti-lung cancer agents in the near future.

Melatonin and endothelial cell-loaded alginate-fibrin hydrogel promoted angiogenesis in rat cryopreserved/thawed ovaries transplanted to the heterotopic sites.

BACKGROUND: Ischemic niche can promote follicular atresia following the transplantation of cryopreserved/thawed ovaries to the heterotopic sites. Thus, the promotion of blood supply is an effective strategy to inhibit/reduce the ischemic damage to ovarian follicles. Here, the angiogenic potential of alginate (Alg) + fibrin (Fib) hydrogel enriched with melatonin (Mel) and CD144+ endothelial cells (ECs) was assessed on encapsulated cryopreserved/thawed ovaries following transplantation to heterotopic sites in rats. METHODS: Alg + Fib hydrogel was fabricated by combining 2% (w/v) sodium Alg, 1% (w/v) Fib, and 5 IU thrombin at a ratio of 4: 2: 1, respectively. The mixture was solidified using 1% CaCl2. Using FTIR, SEM, swelling rate, and biodegradation assay, the physicochemical properties of Alg + Fib hydrogel were evaluated. The EC viability was examined using an MTT assay. Thirty-six adult female rats (aged between 6 and 8 weeks) with a normal estrus cycle were ovariectomized and enrolled in this study. Cryopreserved/thawed ovaries were encapsulated in Alg + Fib hydrogel containing 100 µM Mel + CD144+ ECs (2 × 104 cells/ml) and transplanted into the subcutaneous region. Ovaries were removed after 14 days and the expression of Ang-1, and Ang-2 was monitored using real-time PCR assay. The number of vWF+ and α-SMA+ vessels was assessed using IHC staining. Using Masson's trichrome staining, fibrotic changes were evaluated. RESULTS: FTIR data indicated successful interaction of Alg with Fib in the presence of ionic cross-linker (1% CaCl2). Data confirmed higher biodegradation and swelling rates in Alg + Fib hydrogel compared to the Alg group (p < 0.05). Increased viability was achieved in encapsulated CD144+ ECs compared to the control group (p < 0.05). IF analysis showed the biodistribution of Dil+ ECs within hydrogel two weeks after transplantation. The ratio of Ang-2/Ang-1 was statistically up-regulated in the rats that received Alg + Fib + Mel hydrogel compared to the control-matched groups (p < 0.05). Based on the data, the addition of Mel and CD144+ ECs to Alg + Fib hydrogel reduced fibrotic changes. Along with these changes, the number of vWF+ and α-SMA+ vessels was increased in the presence of Mel and CD144+ ECs. CONCLUSIONS: Co-administration of Alg + Fib with Mel and CD144+ ECs induced angiogenesis toward encapsulated cryopreserved/thawed ovarian transplants, resulting in reduced fibrotic changes.

Epigenetic Modification Factors and microRNAs Network Associated with Differentiation of Embryonic Stem Cells and Induced Pluripotent Stem Cells toward Cardiomyocytes: A Review.

More research is being conducted on myocardial cell treatments utilizing stem cell lines that can develop into cardiomyocytes. All of the forms of cardiac illnesses have shown to be quite amenable to treatments using embryonic (ESCs) and induced pluripotent stem cells (iPSCs). In the present study, we reviewed the differentiation of these cell types into cardiomyocytes from an epigenetic standpoint. We also provided a miRNA network that is devoted to the epigenetic commitment of stem cells toward cardiomyocyte cells and related diseases, such as congenital heart defects, comprehensively. Histone acetylation, methylation, DNA alterations, N6-methyladenosine (m6a) RNA methylation, and cardiac mitochondrial mutations are explored as potential tools for precise stem cell differentiation.

Electrochemical biosensors in exosome analysis; a short journey to the present and future trends in early-stage evaluation of cancers.

The tumor microenvironment consists of a multiplicity of cells such as cancer cells, fibroblasts, endothelial cells, and immune cells within the specific parenchyma. It has been indicated that cancer cells can educate other cells within the tumor niche in a paracrine manner by the release of nano-sized extracellular vesicles namely exosomes (Exo), resulting in accelerated tumor mass growth. It is suggested that exosomal cargo with remarkable information can reflect any changes in metabolic and proteomic profiles in parent tumor cells. Therefore, exosomes can be touted as prognostic, diagnostic, and therapeutic elements with specific biomarkers in patients with different tumor types. Despite the advantages, conventional exosome separation and purification protocols are time-consuming and laborious with low abnormal morphology and purity rate. During the last decades, biosensor-based modalities, as emerging instruments, have been used to detect and analyze Exo in biofluids. Due to suitable specificity, sensitivity, and real-time readout, biosensors became promising approaches for the analysis of Exo in in vitro and in vivo settings. The inherent advantages and superiority of electrochemical biosensors in the determination of tumor grade based on exosomal cargo and profile were also debated. Present and future challenges were also discussed related to the application of electrochemical biosensors in the clinical setting. In this review, the early detection of several cancer types associated with ovaries, breast, brain, colon, lungs, T and B lymphocytes, liver and rare types of cancers were debated in association with released exosomes.

Type 2 diabetes mellitus stimulated pulmonary vascular inflammation and exosome biogenesis in rats.

It has been shown that type 2 Diabetes Mellitus (T2DM) changes the paracrine activity of several cell types. Whether the biogenesis of exosomes is changed during diabetic conditions is the subject of debate. Here, we investigated the effect of T2M on exosome biogenesis in rat pulmonary tissue. Rats received a high-fat diet regime and a single low dose of Streptozocin to mimic the T2DM-like condition. A total of 8 weeks after induction of T2DM, rats were subjected to several analyses. Besides histological examination, vascular cell adhesion molecule 1 (VCAM-1) levels were detected using immunohistochemistry (IHC) staining. Transcription of several genes such as IL-1ß, Alix, and Rab27b was calculated by real-time polymerase chain reaction assay. Using western blot analysis, intracellular CD63 levels were measured. The morphology and exosome secretion activity were assessed using acetylcholinesterase (AChE) assay and scanning electron microscopy, respectively. Histological results exhibited a moderate-to-high rate of interstitial pneumonia with emphysematous changes. IHC staining showed an increased VCAM-1 expression in the diabetic lungs compared with the normal conditions (p < .05). Likewise, we found the induction of IL-1ß, and exosome-related genes Alix and Rab27b under diabetic conditions compared with the control group (p < .05). Along with these changes, protein levels of CD63 and AChE activity were induced upon the initiation of T2DM, indicating accelerated exosome biogenesis. Taken together, current data indicated the induction of exosome biogenesis in rat pulmonary tissue affected by T2DM. It seems that the induction of inflammatory niche is touted as a stimulatory factor to accelerate exosome secretion.

Classical music restored fertility status in rat model of premature ovarian failure.

BACKGROUND: The restorative effect of classical music was assessed on the cyclophosphamide-induced animal model of premature ovarian failure (POF). METHODS: Mozart's piano classical music (K.448) was used for up to 4 and 8 weeks. Rats were exposed to music 6 h every day using a stereo system with a volume of 65-70 dB. Sera and ovarian tissue samples were collected for the evaluation of FSH, LH, and E2 and histopathological examination. At the same time points, samples were taken from the hypothalamus and hippocampus to monitor the expression of Ntrk2, Crh, and Pomc using real-time PCR. Mating trial was performed to evaluate the fertility status of POF rats. RESULTS: Histopathological examination revealed a significant increase (p < 0.05) in the numbers of morphologically normal follicles at all the developmental stages in POF rats after music therapy compared to the POF group (p < 0.05). Music therapy decreased FSH and LH levels to near-to-normal levels conidied with elevation of E2 (p < 0.05). Ntrk2, Crh, and Pomc expressions were down-regulated in POF rats. Music therapy increasaed the expression of Ntrk2 in the hypothalamus of POF rats (p < 0.05). In contrast, Crh and Pomc failed to reach the detection limit before intervention and four weeks after the intervention however, these genes were expressed eight weeks after music therapy. Fertility status was increased (p < 0.05) in terms of litter size in POF rats after being exposed to music compared to the non-treated POF control group (p < 0.05). CONCLUSION: Results showed that music can exert therapeutic effects on POF rats via the alteration of sex-related hormones.

Decellularized bovine ovarian niche restored the function of cumulus and endothelial cells.

OBJECTIVE: Recently, the decellularization technique is introduced as one of the tissue engineering procedures for the treatment of various deficiencies. Here, we aimed to assess the dynamic activity of CCs and HUVECs within decellularized bovine ovarian tissue transplanted subcutaneously in rats. Ovarian tissue was decellularized using a cocktail consisting of different chemicals, and the efficiency of decellularization was assessed using hematoxylin-eosin and DAPI staining. The cell survival was evaluated using an LDH leakage assay. Thereafter, decellularized samples were recellularized using HUVECs and CCs, encapsulated inside alginate (1.2%)-gelatin, (1%) hydrogel, and transplanted subcutaneously to rats. The existence of CD31- and estrogen-positive cells was assessed using immunohistochemistry staining. RESULTS: Bright-field imaging and DAPI staining revealed the lack of nuclei with naive matrix structure in ovarian tissue subjected to decellularization protocol. SEM imaging revealed a normal matrix in decellularized ovaries. LDH assay showed a lack of cytotoxicity for CCs after 7-days compared to the control group. Immunohistochemistry staining showed both CD31- and estrogen-positive cells in CCs + HUVECs compared to the CCs group. CD31 cells appeared with flattened morphology aligned with matrix fibers. The existence of estrogen and CD31 positive cells showed the efficiency of decellularized ovarian tissue to restore cellular function and activity.