A biomimetic multi-layer scaffold with collagen and zinc doped bioglass as a skin-regeneration agent in full-thickness injuries and its effects in vitro and in vivo.

Due to the multilayer structure of skin tissue, the fabrication of a 3-layer scaffold could result in planned dermal regeneration. Herein, polyurethane (PU) and polycaprolactone (PCL), as a function of their mechanical stability and collagen due to its arginine-glycine-aspartic acid sequences, zinc ions because of overcoming the common problems of biological factors were employed. The scaffolds' physical, mechanical, and biological properties were examined by SEM, FTIR, contact angle, mechanical tensile, bacteriocidal efficacy, and hemolysis. Also, after L-929 fibroblast seeding, their biological activity was determined by SEM, DAPI, and MTT assays. Then, the cell-seeded scaffolds were implanted in full-thickness wounds of rats and evaluated by wound closure, histological, and molecular techniques. The in vivo studies showed better wound closure with the composite scaffold containing zinc ions. While its dermal re-organization was retarded in the presence of zinc ions compared to the composite scaffold containing non-doped bioglass. Despite this, the doped composite scaffold indicated better observations with the histological evaluations than the nontreated and bare scaffold groups. Real-time PCR confirmed the higher expression of FGF2 and FGFR genes in rats treated with the zinc-doped composite scaffold. In conclusion, PU/PCL-collagen/PCL-collagen containing the doped or non-doped nanoparticles showed better potential to heal dermal injuries.

Overview of Stem Cell Therapy in Intervertebral Disc Disease: Clinical Perspective.

Intervertebral Disc Degeneration (IDD) is recognized as an aging process, an important and most common pathological condition caused by an imbalance of anabolic and catabolic metabolisms in the Intervertebral Disc (IVD), and leads to changes in the Extracellular Matrix (ECM), impaired metabolic regulation of Nucleus Pulposus (NP), and increased oxidative stress. IDD is mostly associated with pain in the back and neck, which is referred to as a type of disability. Pharmacological and surgical interventions are currently used to treat IDD, but evidence has shown that these interventions do not have the ability to inhibit the progression of IDD and restore IVD function because IVD lacks the intrinsic capacity for regeneration. Thus, therapies that rely on a degenerative cell repair mechanism may be a viable alternative strategy. Biological interventions have been assessed by attempting to regenerate IVD by restoring ECM and cellular function. Over the past decade, stem cell-based therapies have been considered, and promising results have been obtained in various studies. Given this, we reviewed clinical trials and preliminary studies of biological disc repair with a focus on stem cell therapy-based therapies.

Mesenchymal Stem Cell-based Therapy and Female Infertility: Limitations and Advances.

Infertility in women can be caused by various female reproductive diseases such as premature ovarian failure (POF), polycystic ovary syndrome (PCOS), endometriosis and Asherman syndrome that affect couples' quality of life and lead to mental, emotional, and physical problems. In recent years, clinical researchers have sought infertility treatments using new methods that are more effective and noninvasive than the old methods. Today, stem cell-based therapy has been introduced as a promising method and an alternative to the old strategy of infertility treatment. Understanding the main features and functional perspective of mesenchymal stem cells (MSCs) in the future of infertility by physicians is crucial. Mesenchymal stem cells (MSCs) are multipotent stem cells with a high proliferation range, abundant source and multidirectional differentiation potential. They have a high potential for the treatment of injured tissues in regenerative medicine through cell homing, secretion of active factors, and participation in immune regulation. At present, due to fewer ethical restrictions on the use of mesenchymal stem cells compared to embryonic stem cells, more attention has been paid to these cells as a new treatment for gynecological disorders. In this paper, we first review the various type of female reproductive disorders along with their common treatment methods, then we evaluate the recent advances in the application of MSCs in the diseases related to infertility and improve the reproductive health of women worldwide.

Ionizing Radiation Promotes Epithelial-Mesenchymal Transition Phenotype and Stem Cell Marker in The Lung adenocarcinoma: In Vitro and Bioinformatic Studiesc.

OBJECTIVE: Ionizing radiation (IR) is one of the major therapeutic approaches in the non-small cell lung cancer (NSCLC); however, it can paradoxically result in cancer progression likely through promoting epithelial-mesenchymal transition (EMT) and the cancer stem cell phenotype. Therefore, we aimed to determine whether IR promote EMT/CSC and to investigate the clinical relevance of EMT/CSC hallmark genes. MATERIALS AND METHODS: In this experimental and bioinformatic study, A549 cell line was irradiated with a high dosage (6 Gy) or a fractionated regimen (2 Gy/day for 15 fractions). The EMT-related features, including cellular morphology, migratory and invasive capacities were evaluated using scratch assay and transwell migration/invasion assays. The mRNA levels of EMT-related genes (CDH1, CDH2, SNAI1 and TWIST1), stemness-related markers (CD44, PROM1, and ALDH1A1) and the CDH2/CDH1 ratio were evaluated via real-time polymerase chain reaction (PCR). The clinical significance of these genes was assessed in the lung adenocarcinoma (LUAD) samples using online databases. RESULTS: Irradiation resulted in a dramatic elongation of cell shape and enhanced invasion and migration capabilities. These EMT-like alterations were accompanied with enhanced levels of SNAI1, CDH2, TWIST1, CD44, PROM1, and ALDH1A1 as well as an enhanced CDH2/CDH1 ratio. TCGA analysis revealed that, TWIST1, CDH1, PROM1 and CDH2 were upregulated; whereas, CD44, SNAI1 and ALDH1A1 were downregulated. Additionally, correlations between SNAI1-TWIST1, CDH2- TWIST1, CDH2-SNAI1, and ALDH1A1-PROM1 was positive. Kaplan-Meier survival analysis identified lower expression of CDH1, PROM1 and ALDH1A1 and increased expression of CDH2, SNAI1, and TWIST1 as well as CDH2/CDH1 ratio predict overall survival. Additionally, downregulation of ALDH1A1 and upregulation of CDH2, SNAI1 and CTWIST1 could predict a shorter first progression. CONCLUSION: Altogether, these findings demonstrated that IR promotes EMT phenotype and stem cell markers in A549 cell line and these genes could function as diagnostic or prognostic indicators in LUAD samples.

Progress in affinity ligand-functionalized bacterial magnetosome nanoparticles for bio-immunomagnetic separation of HBsAg protein.

Efficient Bio-immunomagnetic separation (BIMS) of recombinant hepatitis B surface antigen (rHBsAg) with high binding capacity was studied using affinity ligand immobilized bacterial magnetosome nanoparticles (Magnetospirillum gryphiswaldense strain MSR-1 bacteria) as an immunomagnetic sorbent. Our results showed immunomagnetic adsorption, acted by affinity interactions with the immobilized monoclonal antibody, offered higher antigen adsorption and desorption capacities as compared with the commercially available immunoaffinity sorbents. Four different ligand densities of the Hep-1 monoclonal antibody were examined during covalent immobilization on Pyridyl Disulfide-functionalized magnetosome nanoparticles for HBsAg immunomagnetic separation. The average of adsorption capacity was measured as 3 mg/ml in optimized immunomagnetic sorbent (1.056 mg rHBsAg/ml immunomagneticsorbent/5.5 mg of total purified protein) and 5mg/ml in immunoaffinity sorbent (0.876 mg rHBsAg/ml immunosorbent/5.5 mg total purified protein during 8 runs. Immunomagnetic sorbent demonstrated ligand leakage levels below 3 ng Mab/Ag rHBsAg during 12 consecutive cycles of immunomagnetic separation (IMS). The results suggest that an immunomagnetic sorbent with a lower ligand density (LD = 3 mg Mab/ml matrix) could be the best substitute for the immunosorbent used in affinity purification of r-HBsAg there are significant differences in the ligand density (98.59% (p-value = 0.0182)), adsorption capacity (97.051% (p-value = 0.01834)), desorption capacity (96.06% (p-value = 0.036)) and recovery (98.97% (p-value = 0.0231)). This study indicates that the immunosorbent approach reduces the cost of purification of Hep-1 protein up to 50% as compared with 5 mg Mab/ml immunoaffinity sorbent, which is currently used in large-scale production. As well, these results demonstrate that bacterial magnetosome nanoparticles (BMs) represent a promising alternative product for the economical and efficient immobilization of proteins and the immunomagnetic separation of Biomolecules, promoting innovation in downstream processing.

Strontium doped bioglass incorporated hydrogel-based scaffold for amplified bone tissue regeneration.

Repairing of large bone injuries is an important problem in bone regeneration field. Thus, developing new therapeutic approaches such as tissue engineering using 3D scaffolds is necessary. Incorporation of some bioactive materials and trace elements can improve scaffold properties. We made chitosan/alginate/strontium-doped bioglass composite scaffolds with optimized properties for bone tissue engineering. Bioglass (BG) and Sr-doped bioglasses (Sr-BG) were synthesized using Sol-Gel method. Alginate-Chitosan (Alg/Cs) scaffold and scaffolds containing different ratio (10%, 20% and 30%) of BG (Alg/Cs/BG10, 20, 30) or Sr-BG (Alg/Cs/Sr-BG10, 20, 30) were fabricated using freeze drying method. Characterization of bioglasses/scaffolds was done using zeta sizer, FTIR, XRD, (FE) SEM and EDS. Also, mechanical strength, antibacterial effect degradation and swelling profile of scaffolds were evaluated. Bone differentiation efficiency and viability of MSCs on scaffolds were determined by Alizarin Red, ALP and MTT methods. Cell toxicity and antibacterial effect of bioglasses were determined using MTT, MIC and MBC methods. Incorporation of BG into Alg/Cs scaffolds amplified biomineralization and mechanical properties along with improved swelling ratio, degradation profile and cell differentiation. Mechanical strength and cell differentiation efficiency of Alg/Cs/BG20 scaffold was considerably higher than scaffolds with lower or higher BG concentrations. Alg/Cs/Sr-BG scaffolds had higher mechanical stability and more differentiation efficiency in comparison with Alg/Cs and Alg/Cs/BG scaffolds. Also, Mechanical strength and cell differentiation efficiency of Alg/Cs/Sr-BG20 scaffold was considerably higher than scaffolds with various Sr-BG concentrations. Biomineralization of Alg/Cs/BG scaffolds slightly was higher than Alg/Cs/Sr-BG scaffolds. Overall, we concluded that Alg/Cs/Sr-BG20 scaffolds are more suitable for repairing bone major injuries.

Expression and characterization of hemagglutinin-neuraminidase protein from Newcastle disease virus in Bacillus subtilis WB800.

BACKGROUND: Newcastle disease virus (NDV) belongs to the genus Avaluvirus and Paramyxoviridae family, and it can cause acute, highly contagious Newcastle disease in poultry. The two proteins, haemagglutinin neuraminidase (HN) and Fusion (F), are the main virulence factor of the virus and play an essential role in immunogenicity against the virus. In most paramyxoviruses, the F protein requires HN protein to fuse the membrane, and HN proteins substantially enhance the viruses' fusion activity. RESULTS: The present study describes the successful cloning and expression of HN protein from NDV in Bacillus subtilis WB800 using the modified shuttle vector pHT43. HN coding sequence was cloned into the pGet II vector. It was then subcloned into the PHT43 shuttle vector and transferred to Escherichia coli for replication. The recombinant plasmid was extracted from E. coli and used to transform B. subtilis by electroporation. After induction of recombinant B. subtilis by IPTG, total cell protein and the protein secreted into the media were analysed through a time course using SDS-PAGE. The expressed HN protein was purified using cation exchange chromatography followed by metal affinity chromatography, using the 6× His epitope introduced at the carboxyl terminus of the recombinant protein. The accuracy of the PHT43-HN construct was confirmed by sequencing and enzymatic digestion. SDS-PAGE results showed that the recombinant HN protein was successfully expressed and secreted into the medium. Moreover, the purified HN protein showed neuraminidase activity with characteristics similar to the indigenous HN NDV protein. B. subtilis is a free endotoxin host that could be a favourite prokaryotic platform for producing the recombinant HN protein. CONCLUSION: The establishment of this expression and purification system has allowed us to explore further the biochemical characteristics of HN protein and obtain material that could be suitable for a new production of NDV candidate vaccine with high immunogenicity.

Molecular evaluation and genetic characterisation of Newcastle disease virus's haemagglutinin-neuraminidase protein isolated from broiler chickens in Iran.

BACKGROUND: Newcastle disease (ND) virus (NDV) is one of the major pathogens in poultry farms that causes severe economic damages to the poultry industry, especially broiler chicken and turkey farms. Despite the endemicity of ND and its many epidemics in the country, the nature of the Iranian strain of the Newcastle virus is still largely unknown. This study aimed to characterise and evaluate NDV isolates obtained from commercial poultry farms in Iran in 2019 through haemagglutinin-neuraminidase (HN) gene sequencing. METHOD: HN gene of each NDV isolate was amplified and sequenced using specific primers followed by phylogenetic analysis of full length of HN gene open reading frame and amino acid (aa) sequence of HN. RESULTS: Phylogenetic analysis of the HN gene showed that the virus is very closely related to genotypes VII and III. Analysis of HN gene nucleotide sequences showed that all isolates encode proteins with a length of 571 aa. CONCLUSION: Results of the present study are useful for a better understanding of molecular epidemiology of indigenous NDV strains and determining important molecular differences between fields and commonly used vaccine strains related to main immunogenic proteins.

Stem cell therapy as a recent advanced approach in male infertility.

Infertility is one of the most common problems in the world that has negative effects on society and infertile people. Among the various causes of infertility, male infertility accounts for almost half of all infertility cases. Despite advances in medicine, current male infertility treatments such as assisted reproductive technology (ART) have not been successful in treating all types of male infertility. Recently, stem cells have been considered as therapeutic targets for many diseases, including infertility, due to their self-renewing and high differentiation. The purpose of this review is to discuss different types of male infertility and the effect of various stem cells against the treatment of male infertility.

Molecular and serological detection and of Toxoplasma gondii in small ruminants of southwest Iran and the potential risks for consumers.

Toxoplasmosis is one of the most important zoonotic diseases with serious health risks for humans, especially for immunodeficient patients, and can lead to abortion in pregnant women worldwide. The oral uptake of sporulated oocysts and/or consumption of undercooked/raw meat of animals infected with Toxoplasma gondii can infect other animals and humans. Heart, liver, and meat tissues of 150 sheep and 150 goats from a slaughterhouse in Ahvaz, Iran, were collected during autumn 2018 and analyzed via polymerase chain reaction (PCR) to detect parasitic DNA in the animal tissues. Moreover, antibodies against T. gondii of 150 sera samples were detected as the targets by in-house enzyme-linked immunosorbent assay (in-house ELISA). A total of 26 (17.3%), 33 (22%), and 48 (32%) of liver, meat, and heart samples in sheep, and a total of 24 (16%), 26 (17.3%), and 36 (24%) of liver, meat, and heart samples in goats, respectively, showed positive PCR results. Besides, the ELISA evaluation of sera samples from 150 sheep and 150 goats resulted in 26 (13.3%) and 16 (10.6%) positive cases, respectively. A significant difference was also found between PCR-positive heart samples and ELISA-positive sera samples of both animal species (p < 0.05), but no significant difference existed between PCR-positive liver samples and ELISA-positive sera samples of both species (p > 0.05). The results of this study confirm the presence of T. gondii in sheep and goats' consumable organs, highlighting the need to avoid consuming raw or uncooked organs of these animal species to prevent human infection with T. gondii.

Cytotoxicity Activity and Druggability Studies of Sigmasterol Isolated from Marine Sponge Dysidea avara Against Oral Epithelial Cancer Cell (KB/C152) and T-Lymphocytic Leukemia Cell Line (Jurkat/ E6-1).

BACKGROUND: Marine sponge is a rich natural resource of many pharmacological compounds and various bioactive anticancer agents are derived from marine organisms like sponges. METHODS: studying the anticancer activity and Drug ability of marine sponge Dysidea avara using Cell lines oral epithelial cancer cell (KB/C152) and T-lymphocytic leukemia cell line (Jurkat/ E6-1). Marine sponge was collected from Persian Gulf. Several analytical techniques have been used to obtain and recognize stigmasterol, including column chromatography, thin layer chromatography, and gas chromatography-mass spectrometry. The PASS Prediction Activity was used to investigate the apoptosis-inducing effect of stigmasterol. The cytotoxic activity of stigmasterol was examined using yellow tetrazolium salt XTT (sodium 2, 3,-bis (2methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium) assay. The stigmasterol were docked within the protein tyrosine kinase (PTKs) (PDB code: 1t46) and epidermal growth factor receptor (EGFRK) (PDB code: 1M17). Also, the pharmacological characteristics of stigmasterol were predicted using PerADME, SwissADME, and Molinspi ration tools. Apoptosis-inducing effect of stigmasterol indicate the stigmasterol in terms of the possibility of apoptosis in cells. RESULTS: The apoptosis inducement results of known stigmasterol were determined by PASS on-line prediction. The compound exhibit potent cytotoxic properties against KB/C152 cell compared to Jurkat/ E6-1 cell. The stigmasterol showed the cytotoxicity effects on KB/C152 and HUT78 with IC50 ranges of 81.18 and 103.03 µg/ml, respectively. Molecular docking showed that, stigmasterol bound stably to the active sites of the protein tyrosine kinase (PTKs) (PDB code: 1t46) and epidermal growth factor receptor (EGFRK) (PDB code: 1M17). CONCLUSION: The compound showed desirable pharmacokinetic properties (ADME). This provided direct evidence of how a prospective anti-cancer agent can be stigmasterol. The preclinical studies paved the way for a potential new compound of anti-cancer.

Synthesis and evaluation of a novel nanosized anionic linear globular dendrimer G2-ciprofloxacin conjugate against prostate cancer.

Prostate cancer is the second most common cancer in the world and the fifth cause of cancer deaths in men. Ciprofloxacin enables the inhabitation of the development of prostate cancer. In this regard, we plan to improve the anticancer effect of ciprofloxacin using the anionic G2 dendrimer in conjunction with ciprofloxacin. In the current study, we measured the size and the zeta potential as well as LC Mass to prove the fact that the conjugation was synthesized correctly. The anticancer activity among three groups including Ciprofloxacin, Ciprofloxacin -G2 dendrimer, and control was measured in vitro. In vitro studies showed that G2 anionic linear-globular polyethylene-glycol-based dendrimer, which conjugated to ciprofloxacin, was able to significantly improve the treatment efficacy over clinical ciprofloxacin alone with respect to proliferation assay. Maximal inhibitory concentration (IC50) was calculated as 200 µ/mL for ciprofloxacin alone and 30µ/mL for ciprofloxacin-G2 dendrimer. In addition, the growth of DU-145 cancerous cells was inhibited by ciprofloxacin-G2 dendrimer conjugate and the number of apoptotic and necrotic cells was increased significantly as evaluated by an annexin V-fluorescein isothiocyanate assay. Ciprofloxacin -G2 dendrimer conjugate was able to increase Bcl-2/Bax ratio in a large scale as compared with the control group and CBL alone. According to the above results, this compound could be considered as a good candidate for functional cancer treatment with low side effects.

Effects of Environmental Conditions on High-Yield Magnetosome Production by Magnetospirillum gryphiswaldense MSR-1

Background: Magnetotactic bacteria are a heterogeneous group of Gram-negative prokaryote cells that produce linear chains of magnetic particles called magnetosomes, intracellular organelles composed of magnetic iron particles. Many important applications have been defined for magnetic nanoparticles in biotechnology, such as cell separation applications, as well as acting as carriers of enzymes, antibodies, or anti-cancer drugs. Since the bacterial growth is difficult and the yield of magnetosome production is low, the application of magnetosome has not been developed on a commercial scale. Methods: Magnetospirillum gryphiswaldense strain MSR-1 was used in a modified current culture medium supplemented by different concentrations of oxygen, iron, carbon, and nitrogen, to increase the yield of magnetosomes. Results: Our improved MSR-1 culture medium increased magnetosome yield, magnetosome number per bacterial cell, magnetic response, and bacterial cell growth yield significantly. The yield of magnetosome increased approximately four times. The optimized culture medium containing 25 mM of Na-pyruvate, 40 mM of NaNO3, 200 µM of ferrous sulfate, and 5-10 ppm of dissolved oxygen (DO) resulted in 186.67 mg of magnetosome per liter of culture medium. The iron uptake increased significantly, and the magnetic response of the bacteria to the magnetic field was higher than threefold as compared to the previously reported procedures. Conclusion: This technique not only decreases the cultivation time but also reduces the production cost. In this modified method, the iron and DO are the major factors affecting the production of magnetosome by M. gryphiswaldense strain MSR-1. However, refining this technique will enable a further yield of magnetosome and cell density.

Structural characterization of recombinant streptokinase following recovery from inclusion bodies using different chemical solubilization treatments.

Circular dichroism (CD) in far-UV region was employed to study the extent of changes occurred in the secondary structures of recombinant streptokinase (rSK), solubilized from inclusion bodies (IBs) by different chemicals and refolded/purified by chromatographic techniques. The secondary structure distribution of rSK, obtained following different chemical solubilization systems, was varied and values in the range of 12.4-14.5% α-helices, 40-51% ß-sheets and 35.5-48.3% turns plus residual structures were found. With reducing the concentration of chemicals during IB solubilization, the content of turns plus random coils was diminished and simultaneously the amounts of α- and ß-sheets were increased. These changes in the secondary structures would lower the hydrophobicity along with the chance of protein aggregation and expose the hydrophilic regions of the protein. Therefore, these alterations in the secondary structures, occurred following efficient IBs solubilization by low concentration of chemicals, could be related to enhancement in rSK biological potency previously observed.

Leishmaniasis in humans: drug or vaccine therapy?

Leishmania is an obligate intracellular pathogen that invades phagocytic host cells. Approximately 30 different species of Phlebotomine sand flies can transmit this parasite either anthroponotically or zoonotically through their bites. Leishmaniasis affects poor people living around the Mediterranean Basin, East Africa, the Americas, and Southeast Asia. Affected regions are often remote and unstable, with limited resources for treating this disease. Leishmaniasis has been reported as one of the most dangerous neglected tropical diseases, second only to malaria in parasitic causes of death. People can carry some species of Leishmania for long periods without becoming ill, and symptoms depend on the form of the disease. There are many drugs and candidate vaccines available to treat leishmaniasis. For instance, antiparasitic drugs, such as amphotericin B (AmBisome), are a treatment of choice for leishmaniasis depending on the type of the disease. Despite the availability of different treatment approaches to treat leishmaniasis, therapeutic tools are not adequate to eradicate this infection. In the meantime, drug therapy has been limited because of adverse side effects and unsuccessful vaccine preparation. However, it can immediately make infections inactive. According to other studies, vaccination cannot eradicate leishmaniasis. There is no perfect vaccine or suitable drug to eradicate leishmaniasis completely. So far, no vaccine or drug has been provided to induce long-term protection and ensure effective immunity against leishmaniasis. Therefore, it is necessary that intensive research should be performed in drug and vaccine fields to achieve certain results.

In vitro physical and biological characterization of biodegradable elastic polyurethane containing ferulic acid for small-caliber vascular grafts.

Demand for small diameter vascular grafts is growing. The main limitations of these grafts include induced thrombotic events, lack of in situ endothelialization, intimal hyperplasia and poor mechanical properties which impair the graft patency rate in long-term applications. Most anti-thrombotic modification methods currently in use usually conflict with the formation of an endothelial cell monolayer on the grafts. Here, we synthesized a novel biodegradable poly(ether ester urethane)urea elastomer (PEEUU) using poly(ethylene glycol) and poly(diethylene glycol adipate) as soft segments. To improve hemocompatibility, synthesized PEEUU was blended with ferulic acid (FA). Scanning electron microscopy, water contact angle measurement, and tensile testing were used to characterize the scaffolds. The PEEUU and PEEUU-FA scaffolds revealed appropriate mechanical properties, with tensile strengths and strains similar to a coronary artery. In vitro assay demonstrated that the release of FA from the scaffold is in a sustained manner. Hemocompatibility tests indicated that the PEEUU-FA sample induced lower platelet adhesion compared to the PEEUU sample. Reductions in hemolysis and fibrinogen adsorption were detected on the PEEUU-FA sample. Cell studies showed that PEEUU-FA supported the adhesion, expansion and proliferation of endothelial cells. The cells maintained an endothelial cell phenotype through the expression of the endothelial cell marker CD31. The results revealed that the new PEEUU modified with FA can be considered as a promising candidate for vascular applications with enhanced blood compatibility and vascular cell-compatibility.

Injectable natural polymer compound for tissue engineering of intervertebral disc: In vitro study.

Intervertebral disc degeneration is recognized to be the leading cause for chronic low-back pain. Injectable hydrogel is one of the great interests for tissue engineering and cell encapsulation specially for intervertebral (IVD) affecting rate of regeneration success, in this study we assessed viscoelastic properties of a Chitosan-ß glycerophosphate-hyaluronic acid, Chondroitin-6-sulfate, type 2 of Collagen, gelatin, fibroin silk (Ch-ß-GP-HA-CS-Col-Ge-FS) hydrogel which was named as NP hydrogel that is natural extracellular matrix of IVD. Chitosan-based hydrogel was made in the ratio of 1.5%: 7%: 1%:1%:1%-1.5%-1% (Ch: ß-GP: HA-CS-Col-Ge-FS). Gelation time and other rheological properties were studied using amplitude sweep and frequency sweep tests. Also, the cytotoxicity of the hydrogel invitro assessed by MTT and trypan blue tests. Morphology of the hydrogel and attachment of NP cells were evaluated by SEM. Our result showed that NP hydrogel in 4°C is an injectable transparent solution. It started gelation in 37°C after about 30min. Gelation temperature of NP hydrogel was 37°C. Storage modulus (G') of this hydrogel at 37°C was almost constant over a wide range of strain. MTT and trypan blue tests showed hydrogel was cytocompatible. The obtained results suggest that this hydrogel would be a natural and cytocompatible choice as an injectable scaffold for using in vivo study of IVD regeneration.

Apparent histological changes of adipocytes after treatment with CL 316,243, a ß-3-adrenergic receptor agonist.

BACKGROUND AND OBJECTIVES: The objective of this experiment was to study the effect of CL 316,243 (CL) (a highly selective ß3-adrenergic receptor agonist) on cellular changes occurring in retroperitoneal white adipose tissue (RWAT) of lean and obese rats. METHODS: Ten-month-old lean and obese Zucker rats were implanted subcutaneously with osmotic mini-pumps, infusing either saline or CL (1 mg/kg body weight/day) for 4 weeks. RESULTS: There was no effect of CL on food intake. However, the resting metabolic rate in lean and obese rats increased by 55% and 96% per rat, respectively. Total RWAT weight decreased in both lean and obese rats under influence of CL treatment by 65% and 38%, respectively. Total body weight and body fat were lower in CL treated rats. Detection of uncoupling protein 1 (UCP1) in RWAT was confirmed qualitatively by both immunohistochemistry and immunofluorescence using a rabbit anti rat UCP1 antibody which showed the appearance of a marked increase of this protein in the adipose tissue. Stained semi-thin sections (0.5 µm) also demonstrated abundant nuclei in multilocular adipocytes, in endothelial cells associated with the vasculature, and in interstitial cells. In CL-treated obese rats, a clustering of several multilocular cells around the periphery of a white adipocyte was seen. CONCLUSION: These results indicate that treatment of both lean and obese Zucker rats with CL induces extensive remodeling of RWAT that includes shrinkage of white adipose tissue, appearance of abundant multilocular cells in RWAT together with the appearance of a marked increase of UCP, preferentially in lean rats.

Synthesis and characterization of a novel chemically designed (Globo)3-DTPA-KLH antigen.

In recent years, many experiments have been conducted for the production and evaluation of anticancer glycoconjugated vaccines in developed countries and many achievements have been accomplished with Globo H derivatives. In the current experiment, a new chemically designed triplicate version of (Globo H)3-diethylenetriamine pentaacetic acid (DTPA)-KLH antigen was synthesized and characterized. Immunization with (Globo H)3-DTPA-KLH, a hexasaccharide that is a member of a family of antigenic carbohydrates that are highly expressed in various types of cancers conjugated with DTPA and KLH protein, induced a high level of antibody titer along with an elevated level of IL-4 in mice. Treatment of tumors with the collected sera from immunized mice decreased the tumor size in nude mice as well. None of the immunized mice illustrated any sign of tumor growth after injection of MCF-7 cells compared to the control animals. These findings, based on the newly presented structure of the Globo H antigen, lend exciting and promising evidence for clinical advancement in the development of a therapeutic vaccine in the future.

Evaluation of the proliferation and viability rates of nucleus pulposus cells of human intervertebral disk in fabricated chitosan-gelatin scaffolds by freeze drying and freeze gelation methods.

BACKGROUND: Low back pain is one of the most significant musculoskeletal diseases of our time. Intervertebral disk herniation and central degeneration of the disk are two major reasons for low back pain, which occur because of structural impairment of the disk. The reduction of cell count and extracellular matrix, especially in the nucleus pulposus, causes disk degeneration. Different scaffolds have been used for tissue repairing and regeneration of the intervertebral disk in tissue engineering. Various methods are used for fabrication of the porosity scaffolds in tissue engineering. The freeze drying method has disadvantages such as: It is time consuming, needs high energy, and so on. The freeze-gelation method can save a great deal of time and energy, and large-sized porous scaffolds can be fabricated by this method. In this study, proliferation of the nucleus pulposus (NP) cells of the human intervertebral disk are compromised in the fabricated Chitosan-gelatin scaffolds by freeze drying and freeze gelation methods. MATERIALS AND METHODS: The cells were obtained from the nucleus pulposus by collagenase enzymatic hydrolysis. They were obtained from patients who were undergoing open surgery for discectomy in the Isfahan Alzahra Hospital. Chitosan was blended with gelatin. Chitosan polymer, solution after freezing at -80°C, was immersed in sodium hydroxide (NaOH) solution. The cellular suspension was transferred to each scaffold and cultured in plate for 14 days. Cell viability and proliferation were investigated by Trypan blue and MTT assays. RESULTS: The MTT and Trypan blue assays demonstrated that cell viability and the mean of the cell number showed a significant difference between three and fourteen days, in both scaffolds. Accordingly, there was a significantly decrease in the fabricated chitosan-gelatin scaffold by the freeze-drying method. CONCLUSION: The fabricated chitosan-gelatin scaffold by the freeze-gelation method prepared a better condition for proliferation of NP cells when compared with the fabricated chitosan-gelatin scaffold by the freeze drying method.