Global landscape of SARS-CoV-2 mutations and conserved regions.

BACKGROUND: At the end of December 2019, a novel strain of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) disease (COVID-19) has been identified in Wuhan, a central city in China, and then spread to every corner of the globe. As of October 8, 2022, the total number of COVID-19 cases had reached over 621 million worldwide, with more than 6.56 million confirmed deaths. Since SARS-CoV-2 genome sequences change due to mutation and recombination, it is pivotal to surveil emerging variants and monitor changes for improving pandemic management. METHODS: 10,287,271 SARS-CoV-2 genome sequence samples were downloaded in FASTA format from the GISAID databases from February 24, 2020, to April 2022. Python programming language (version 3.8.0) software was utilized to process FASTA files to identify variants and sequence conservation. The NCBI RefSeq SARS-CoV-2 genome (accession no. NC_045512.2) was considered as the reference sequence. RESULTS: Six mutations had more than 50% frequency in global SARS-CoV-2. These mutations include the P323L (99.3%) in NSP12, D614G (97.6) in S, the T492I (70.4) in NSP4, R203M (62.8%) in N, T60A (61.4%) in Orf9b, and P1228L (50.0%) in NSP3. In the SARS-CoV-2 genome, no mutation was observed in more than 90% of nsp11, nsp7, nsp10, nsp9, nsp8, and nsp16 regions. On the other hand, N, nsp3, S, nsp4, nsp12, and M had the maximum rate of mutations. In the S protein, the highest mutation frequency was observed in aa 508-635(0.77%) and aa 381-508 (0.43%). The highest frequency of mutation was observed in aa 66-88 (2.19%), aa 7-14, and aa 164-246 (2.92%) in M, E, and N proteins, respectively. CONCLUSION: Therefore, monitoring SARS-CoV-2 proteomic changes and detecting hot spots mutations and conserved regions could be applied to improve the SARS-CoV-2 diagnostic efficiency and design safe and effective vaccines against emerging variants.

SARS-CoV-2 Non-structural protein 1(NSP1) mutation virulence and natural selection: Evolutionary trends in the six continents.

OBJECTIVE: Rapid transmission and reproduction of RNA viruses prepare conducive conditions to have a high rate of mutations in their genetic sequence. The viral mutations make adapt the severe acute respiratory syndrome coronavirus 2 in the host environment and help the evolution of the virus then also caused a high mortality rate by the virus that threatens worldwide health. Mutations and adaptation help the virus to escape confrontations that are done against it. METHODS: In the present study, we analyzed 6,510,947 sequences of non-structural protein 1 as one of the conserved regions of the virus to find out frequent mutations and substitute amino acids in comparison with the wild type. NSP1 mutations rate divided into continents were different. RESULTS: Based on this continental categorization, E87D in global vision and also in Europe notably increased. The E87D mutation has signed up to January 2022 as the first frequent mutation observed. The remarkable mutations, H110Y and R24C have the second and third frequencies, respectively. CONCLUSION: According to the important role of non-structural protein 1 on the host mRNA translation, developing drug design against the protein could be so hopeful to find more effective ways the control and then treatment of the global pandemic coronavirus disease 2019.

The First Geographic Identification by Country of Sustainable Mutations of SARS-COV2 Sequence Samples: Worldwide Natural Selection Trends.

The high mutation rates of RNA viruses, coupled with short generation times and large population sizes, allow viruses to evolve rapidly and adapt to the host environment. The rapidity of viral mutation also causes problems in developing successful vaccines and antiviral drugs. With the spread of SARS-CoV-2 worldwide, thousands of mutations have been identified, some of which have relatively high incidences, but their potential impacts on virus characteristics remain unknown. The present study analyzed mutation patterns, SARS-CoV-2 AASs retrieved from the GISAID database containing 10,500,000 samples. Python 3.8.0 programming language was utilized to pre-process FASTA data, align to the reference sequence, and analyze the sequences. Upon completion, all mutations discovered were categorized based on geographical regions and dates. The most stable mutations were found in nsp1(8% S135R), nsp12(99.3% P323L), nsp16 (1.2% R216C), envelope (30.6% T9I), spike (97.6% D614G), and Orf8 (3.5% S24L), and were identified in the United States on April 3, 2020, and England, Gibraltar, and, New Zealand, on January 1, 2020, respectively. The study of mutations is the key to improving understanding of the function of the SARS-CoV-2, and recent information on mutations helps provide strategic planning for the prevention and treatment of this disease. Viral mutation studies could improve the development of vaccines, antiviral drugs, and diagnostic assays designed with high accuracy, specifically useful during pandemics. This knowledge helps to be one step ahead of new emergence variants.

Review of Bioprinting in Regenerative Medicine: Naturally Derived Bioinks and Stem Cells.

Regenerative medicine offers the potential to repair or substitute defective tissues by constructing active tissues to address the scarcity and demands for transplantation. The method of forming 3D constructs made up of biomaterials, cells, and biomolecules is called bioprinting. Bioprinting of stem cells provides the ability to reliably recreate tissues, organs, and microenvironments to be used in regenerative medicine. 3D bioprinting is a technique that uses several biomaterials and cells to tailor a structure with clinically relevant geometries and sizes. This technique's promise is demonstrated by 3D bioprinted tissues, including skin, bone, cartilage, and cardiovascular, corneal, hepatic, and adipose tissues. Several bioprinting methods have been combined with stem cells to effectively produce tissue models, including adult stem cells, embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and differentiation techniques. In this review, technological challenges of printed stem cells using prevalent naturally derived bioinks (e.g., carbohydrate polymers and protein-based polymers, peptides, and decellularized extracellular matrix), recent advancements, leading companies, and clinical trials in the field of 3D bioprinting are delineated.

Update review on five top clinical applications of human amniotic membrane in regenerative medicine.

Due to the increasing number of studies performed in the field of regenerative medicine during the last two decades, more analytic studies are still needed to clarify the future prospect of this area of science. The main aim of this research was to review the clinical applications of human Amniotic membrane in the field of regenerative medicine critically. Furthermore, in the light of increasing numbers of available products derived from amniotic membrane, we aimed look in depth to see whether regenerative medicine research strategies have a place in the clinical setting. More specifically, in the present study, we attempted to provide insight on developing the new indication for more research and in the next step, for market leaders companies to expand cost-effectiveness of new derived AM products. 20 companies or distributers have offered some commercial products in this field. Survey on more than 90 clinical trials in last five years showed dermatology (and more specific wound healing), orthopedic, and ophthalmology are heavily biased toward multibillion dollar industry. Moreover, urology and dentistry with fewer numbers of clinical data in comparison with the above-mentioned areas, currently are in the path of translation (especially dentistry). In addition, otolaryngology and oncology with the lowest number showed more potential of research thorough understanding the properties that will help guiding the use of AM-derived products in these two areas in future. More than 50% of clinical studies were done or are developing in USA, which have the biggest share in market products. Subsequently, China, Egypt, India, Iran, and Germany with the ongoing clinical trials in different phases may have more approved products in near future.

Engineered skin graft with stromal vascular fraction cells encapsulated in fibrin-collagen hydrogel: A clinical study for diabetic wound healing.

Despite the abundance of skin substitutes in the worldwide market, major hurdles in developing more complex tissues include the addition of skin appendages and vascular networks as the most important structure. The aim of this research was a clinical feasibility study of a novel prevascularized skin grafts containing the dermal and epidermal layer using the adipose stromal vascular fraction (SVF)-derived endothelial cell population for vascular network regeneration. Herein, we characterized hydrogel with emphasis on biological compatibility and cell proliferation, migration, and vitality. The therapeutic potential of the prevascularized hydrogel transplanted on five human subjects as an intervention group with diabetic wounds was compared with nonvascularized skin grafts as the control on five patients. Wound planimetric and biometric analysis was performed using a Mann-Whitney nonparametric t-test (p ≤ .05). The fibrin-collagen hydrogel was suitable for skin organotypic cell culture. There was a significant (p ≤ .05) increased in skin thickness and density in the vascular beds of the hypodermis measured with skin scanner compared with that in the control group. No significant macroscopic differences were observed between the intervention and control groups (p ≤ .05). In summary, we report for the first time the use of autologous dermal-epidermal skin grafts with intrinsic vascular plexus in a clinical feasibility study. The preliminary data showed that SVF-based full-thickness skin grafts are safe and accelerate the wound healing process. The next stage of the study is a full-scale randomized clinical trial for the treatment of patients with chronic wounds.

A gene-based anti-angiogenesis therapy as a novel strategy for cancer treatment.

Angiogenesis-targeted therapy of cancer is considered a promising strategy for therapeutic management of cancer progression. Over the last two decades, a few anti-angiogenesis monoclonal antibodies (mAbs) blocking VEGF signaling have been developed and approved by the FDA. The most widely used anti-angiogenesis drug is bevacizumab which binds VEGFA and prevents its interaction with VEGF receptor leading to suppression of angiogenesis. Despite the remarkable success in development of angiogenesis inhibitory mAbs, their clinical application is limited by the high-cost of mAbs-based regimen which includes multiple doses of mAbs due to their short biological half-life. Antibody gene therapy is an alternative system of antibody production. In this study, we have developed a gene-based anti-VEGF mAb system which is expected to produce a high concentration of anti-VEGFA mAb upon a single administration in cancer patients. The full-length cDNA bevacizumab light and heavy chains joint with T2A sequence were cloned in pCDH lentivirus vector. The lentiviral particles expressing bevacizumab was produced in HEK-293T cells. Recombinant lentiviral particles containing bevacizumab (rLV-bev) efficiently transduced HEK-293cells and produced functional bevacizumab mAb. Bevacizumab expression in the transduced cell was assessed by qRT-PCR and western blot at both the mRNA and protein level, respectively. The functionality of the recombinant bevacizumab was confirmed using the tube formation assay in the co-culture system of endothelial cells and HT-29cells transduced with rLV-bev viral particles. Our results show that rLV-bev gene therapy can be useful for angiogenesis-targeted therapy of cancer.

Skin regenerative medicine advancements in the Islamic Republic of Iran: a concise review.

In the last decade, the Islamic Republic of Iran has witnessed significant improvement and growth in the field of interdisciplinary medicine and in its translation to patients, including the field of cell and stem cell therapy. The main aim of this report is to highlight various advances in regenerative medicine for skin and dermatology using stem cell technology, and its translation to clinic in the past two decades, in Iranian academic centers, clinical institutes and hospitals. While there have been numerous positive advances in clinical outcomes reported in Iran, there is no comparative analytical information on these studies. Here we present a historical overview of the progress and key advancements seen in skin regeneration in this country, review the research frameworks, regulatory approach and pathways and offer perspectives for the future.

Regenerative Medicine Applications in Wound Care.

BACKGROUND: During the last two decades, a number of studies have been carried out on the application of regenerative medicine in the field of dermatology. OBJECTIVE: The aim of this research was to critically review the application of regenerative medicine in the field of dermatology. The next aim was to look in depth to see whether regenerative medicine strategies have a place in the future of wound healing in a clinical setting. More specifically, to see if these strategies would apply for burns and non-healing diabetic wounds. RESULTS: Billions of dollars have been spent worldwide on research in wound treatment and skin regeneration. Although a high number of clinical trials show promising results, there is still no commercially available treatment for use. In addition, the outcome data from the clinical trials, taking place throughout the world, are not published in a standardized manner. Standardization within clinical trials is required for: protocols, outcome, endpoint values, and length of follow-up. The lack of standardization makes it much more difficult to compare the data collected and the different types of treatment. CONCLUSION: Despite several promising results from research and early phase clinical studies, the treatment for wounds as well as skin regeneration is still considered as an unmet clinical need. However, in the past three years, more promising research has been approaching clinical trials; this could be the solution that clinicians have been waiting for. This is a multibillion dollar industry for which there should be enough incentive for researchers and industry to seek the solution.

Relationship between erb-B2 mRNA Expression in Blood and Tissue of Invasive Ductal Carcinoma Breast Cancer Patients and Clinicopathological Characteristics of the Tumors.

Molecular detection methods such as RT-PCR for detecting breast cancer-associated gene expression in the peripheral blood have the potential to modify breast cancer (BC) staging and therapy. In this regard, we evaluated the potential of erb-B2 molecular marker in BC detection and analyzed the expression of erb-B2 mRNA in the peripheral blood and fresh tissue samples of 50 pretreated female BC patients and 50 healthy females by reverse transcription-PCR (RT-PCR) method. We also assessed the correlation of erb-B2 mRNA marker positivity in peripheral blood and tumor tissue samples with clinical and pathological factors in BC patients in order to evaluate its prognostic value. It was shown that there is a significant difference between healthy females and BC patients with expression of the erb-B2 molecular marker (p<0.01). A significant difference between the expression of erb-B2 in the peripheral blood and tissue samples of BC patients (p<0.01) and the frequency of circulating erb-B2 mRNA expression in peripheral blood and in tissue was detected by RT-PCR. No correlation was found between erb-B2 mRNA expression in blood or tumor tissue samples and lymph node, tumor grade, tumor stage, tumor size, patient's age, ki67, estrogen receptor (ER), progesterone receptor (PGR), P53, and HER-2 status. However, in a small subset of 31 BC patients we found that expression of erb-B2 in peripheral blood or in both peripheral blood and tumor tissue was directly correlated with lympho-vascular invasion and perineural invasion as poor prognostic features. The highest rates of erb-B2 expression in peripheral blood or tumor tissue were in the ER and PR negative and HER-2 positive group. This study suggests that the application of the RT-PCR and immunohistochemical methods for erb-B2 molecular marker detection would provide a higher detection rate, especially in early stage BC.