Melanoma Cell Adhesion Molecule (CD 146) in Endometrial Physiology and Disorder.

The human endometrium, the innermost lining of the uterus, is the anatomic prerequisite for pregnancy. It is the only dynamic tissue that undergoes more than 400 cycles of regeneration throughout the reproductive life of women. Key to this function are endometrial stem cells as well as cell adhesion molecules. Melanoma cell adhesion molecule (MCAM/CD146/MUC18) is a membrane glycoprotein of the mucin family and a key cell adhesion protein, highly expressed by endometrial cells. CD146 is a significant molecule pivotal in endometrial physiology, assisting tissue regeneration and angiogenesis. Endometrium also acts as a culprit in causing several endometrial dysfunctions, such as endometriosis, endometrial hyperplasia, and endometrial carcinoma, due to interrupted molecular and functional mechanisms. Though most of the endometrial dysfunctions arise as a result of endocrine disturbance, it has a major pathological role associated with angiogenesis. It has already been proven that CD146 is a potential marker for the diagnosis of angiogenic dysfunctions and malignancy, including endometrial cancer. However, its mechanistic role in causing the pathology is a mystery. This chapter explores the role of CD146 in normal and pathological endometrial conditions and the therapeutic implications of CD146.

Recellularization of Acellular Xeno Kidney Scaffold: An In Vivo Method to Generate Bioartificial Kidney.

A significant hurdle for kidney tissue engineering is reproducing the complex three-dimensional structure of the kidney. In our study, a stepwise approach of generating a reproducible Xeno kidney scaffold from a goat kidney is described, which can be implanted and recellularized by host cells. We have proposed a combination of sodium dodecyl sulfate and Triton-X-100-based protocol to generate a reproducible Xeno kidney scaffold, which was then analyzed by histology, DNA quantification, SEM, and renal angiography. Further, a small portion from the cortico-medullar region of the acellular scaffold was implanted in the rat's kidney subcapsular pocket for a period of 1 month, to check the recruitment of host cells into the scaffold. Post implantation, the extracellular matrix of the scaffold was well preserved and it did not induce any damage or inflammation in the native kidney. Implantation of the Xeno scaffold resulted in apparent early vascularization which helped in the recruitment of the host cells, which was characterized by histology, immunohistochemistry, and scanning electron microscopy. Implanted Xeno scaffold showed AQP-1, Nephrin, α-SMA, and VEGF expression in proximal tubules and renal glomerulus. Importantly, Ki-67 and WTAP-expressing cells were also observed near proximal tubules suggesting a high level of proliferation in the scaffold. Thus, showing the potential of Xeno kidney development that can be recellularized by the host cell to engineer into a functional kidney.

Pathophysiology of Spinal Cord Injury and Tissue Engineering Approach for Its Neuronal Regeneration: Current Status and Future Prospects.

A spinal cord injury (SCI) is a very debilitating condition causing loss of sensory and motor function as well as multiple organ failures. Current therapeutic options like surgery and pharmacotherapy show positive results but are incapable of providing a complete cure for chronic SCI symptoms. Tissue engineering, including neuroprotective or growth factors, stem cells, and biomaterial scaffolds, grabs attention because of their potential for regeneration and ability to bridge the gap in the injured spinal cord (SC). Preclinical studies with tissue engineering showed functional recovery and neurorestorative effects. Few clinical trials show the safety and efficacy of the tissue engineering approach. However, more studies should be carried out for potential treatment modalities. In this review, we summarize the pathophysiology of SCI and its current treatment modalities, including surgical, pharmacological, and tissue engineering approaches following SCI in preclinical and clinical phases.

Activated platelet-rich plasma accelerate endometrial regeneration and improve pregnancy outcomes in murine model of disturbed endometrium.

Platelet Rich Plasma (PRP) contains high concentrations of growth factors, therefore, PRP activation results in their release, stimulating the process of healing and regeneration. The study was conducted to check whether activated platelet-rich plasma (aPRP) treatment can improve regeneration of the endometrium in an experimental model of ethanol-induced disturbed endometrium. Seventy-two female Wistar rats were randomly assigned into the control group, disturbed endometrium (DE) group and aPRP treated group. Activation of PRP was performed by adding thrombin. All the animals were sacrificed on day 1, day 3, day 6 and day 9 and samples were taken from the miduterine horn. Quantification of Cytokine and chemokine profiles of activated and non-activated PRP for CCL2, TNF- α, IL-1ß, CXCL8, CXCL10, IL2, IL4, IL-6 IL-10, IL-12, IL-17A, TGF- ß, IFN-γ was carried out. Functional and structural recovery of the endometrium was analyzed by hematoxylin-eosin (HE) and immunohistochemical (IHC) analyses. HE confirmed proliferated epithelial lining and stromal reconstruction with decreased fibrosis in PRP treated group compared to the DE group. Epithelial thickness in aPRP treated on day 1, day 3, day 6 and day 9 revealed an significant increase (p ≤ 0.05). Significantly stronger IHC expression of alpha smooth muscle actin, Cytokeratin 18, Cytokeratin 19, Connexin-40, E-Cadherin, Claudin-1, Zona Occludin-1was found in the aPRP treated group compared to the DE group. Furthermore, aPRP treatment was associated with birth of live pups. Our results suggest that intrauterine administration of aPRP stimulated and accelerated the regeneration of endometrium in the murine model of disturbed endometrium.

Tissue engineering of human ear pinna.

Auricular deformities (Microtia) can cause physical, social as well as psychological impacts on a patient's wellbeing. Biofabrication of a complex structure such as ear pinna is not precise with currently available techniques. These limitations can be overcome with the help of tissue engineering. In this article, the authors presented molding and three dimensional (3D) printing to generate a flexible, human size ear pinna. The decellularization of goat ear cartilage protocol and bioink alkaline digestion protocol was followed to yield complete removal of all cellular components without changing the properties of the Extra Cellular Matrix (ECM). Decellularized scaffold used in molding technology and 3D printing technology Computer-Aided Design /Stereolithography (CAD/STL) uses bioink to construct the patient-specific ear. In vivo biocompatibility of the both ear pinnae showed demonstrable recellularization. Histology and scanning electron microscopy analysis revealed the recellularization of cartilage-specific cells and the development of ECM in molded and 3D printed ear pinna after transplantation. Both the techniques provided ideal results for mechanical properties such as elasticity. Vascular Associated Protein expression revealed specific vasculogenic pattern (angiogenesis) in transplanted molded pinna. Chondrocyte specific progenitor cells express CD90+ which highlighted newly developed chondrocytes in both the grafts which indicated that the xenograft was accepted by the rat. Transplantation of molded as well as 3D ear pinna was successful in an animal model and can be available for clinical treatments as a medical object to cure auricular deformities.

Tissue engineered human ear pinna derived from decellularized goat ear cartilage: clinically useful and biocompatible auricle construct.

Surgery of the entire ear pinna even today presents a challenge to reconstructive surgeons, in the absence of a universally acceptable, quality construct for clinical use. In this article, the authors present a technique to generate a flexible, human size ear with the aim to meet this limitation for ear reconstructive surgeries. The construct was engineered by using a decellularized goat ear cartilage. This was characterized by hematoxylin-eosin (H/E), diamidino-2-phenylindole (DAPI), Masson's trichrome (MT), Alcian Blue (AB) staining and Scanning Electron Microscopy (SEM) for extracellular matrix (ECM) analysis. The decellularization protocol followed yielded complete removal of all cellular components without changing the properties of the ECM. In vivo biocompatibility of the ear pinna showed demonstrable recellularization. Recellularization was tracked using HE, DAPI, MT, AB staining, toluidine staining, SEM, vascular-associated protein (VAP) and CD90+ expressing cells. VAP expression revealed specific vasculogenic pattern (angiogenesis). CD90+ expression reflected the presence of the stromal cell. The graft maintained the properties of ECM and displayed chondrocyte recruitment. In summary, the decellularized goat ear pinna (cartilage) exhibited xenograft biocompatibility, stable mechanical properties and in vivo chondrocyte recruitment. Subsequently developed tissue-engineered ear pinna offer potential for cartilage flexibility and individualization of ear shape and size for clinical application.

Decellularized amnion scaffold with activated PRP: a new paradigm dressing material for burn wound healing.

Direct application of amnion has greater risk of immunological rejection and infection. Decellularization is an effective method to lower the risk of immune complications and infections. The bioreactor assembly with multiple cassettes was designed for decellurization of multiple amnions with different cell types simultaneously in single run. A detergent-based protocol was modified to remove all cellular components from amnion and diminish the DNA content to render it non-immunogenic. Amnion (n = 10) were treated with 2% sodium dodecyl sulphate (SDS), 5% dimethyl sulfoxide (DMSO) and 2% sodium deoxycholeate (SD). Decellularized amnion samples were analyzed by haematoxylin-eosin staining (HE), Alcian blue pH 1 (AB-pH-1), 4,6-diamnionidino-2-phenylindol (DAPI), Massion's trichrome stain, DNA quantification, mechanical testing and scanning electron microscopy (SEM). Histological analysis showed complete removal of cellular components and the histoarchitecture of scaffold remained intact. Amnion scaffold activated with platelet rich plasma (PRP) and calcium chloride composition supported better adherence to the wound than amnion alone. Only single application showed good healing. In vivo assessment of activated amnion revealed stable dressing. It has good promising outcome. At day 7, histologically the wounds treated with activated amnion were almost closed without scarring and showed well differentiated epidermis, proliferation of keratinocytes, hair follicles and basement membrane as compared to controls and silver nitrate gel dressings in a mouse (Mus musculus). Cryopreservation had no adverse effect on the mechanical properties of the amnion scaffold. Cryopreservation of decellularized amnion by Dulbecco's modified eagle medium (DMEM) was expected to prepare off-the-shelf skin substitutes and preserve them to be immediately available upon request of patients' needs.

TEMPORARY REMOVAL: CD271 identifies functional human hepatic stellate cells, which localize in peri-sinusoidal and portal areas in liver after partial hepatectomy.

BACKGROUND AIMS: Hepatic stellate cells (HSCs) are liver-resident mesenchymal cells involved in essential processes in the liver. However, knowledge concerning these cells in human livers is limited because of the lack of a simple isolation method. METHODS: We isolated fetal and adult human liver cells by immunomagnetic beads coated with antibodies to a mesenchymal stromal cell marker (CD271) to enrich a population of HSCs. The cells were characterized by cell cultivation, immunocytochemistry, flow cytometry, reverse-transcription polymerase chain reaction and immunohistochemistry. Cells were injected into nude mice after partial hepatectomy to study in vivo localization of the cells. RESULTS: In vitro, CD271(+) cells were lipid-containing cells expressing several HSC markers: the glial fibrillary acidic protein, desmin, vimentin and α-smooth muscle actin but negative for CK8, albumin and hepatocyte antigen. The cells produced several inflammatory cytokines such as interleukin (IL)-6, IL-1A, IL-1B and IL-8 and matrix metalloproteinases MMP-1 and MMP-3 and inhibitors TIMP-1 and TIMP-2. In vivo, fetal CD271(+) cells were found in the peri-sinusoidal space and around portal vessels, whereas adult CD271(+) cells were found mainly in the portal connective tissue and in the walls of the portal vessels, which co-localized with α-smooth muscle actin or desmin. CD271(-) cells did not show this pattern of distribution in the liver parenchyma. CONCLUSIONS: The described protocol establishes a method for isolation of mesenchymal cell precursors for hepatic stellate cells, portal fibroblasts and vascular smooth muscle cells. These cells provide a novel culture system to study human hepatic fibrogenesis, gene expression and transcription factors controlling HSC regulation.

Phenotypic and in vivo functional characterization of immortalized human fetal liver cells.

We report the establishment and characterization of immortalized human fetal liver progenitor cells by expression of the Simian virus 40 large T (SV40 LT) antigen. Well-characterized cells at various passages were transplanted into nude mice with acute liver injury and tested for functional capacity. The SV40LT antigen-immortalized fetal liver cells showed a morphology similar to primary cells. Cultured cells demonstrated stable phenotypic expression in various passages, of hepatic markers such as albumin, CK 8, CK18, transcription factors HNF-4α and HNF-1α and CYP3A/7. The cells did not stain for any of the tested cancer-associated markers. Albumin, HNF-4α and CYP3A7 expression was confirmed by reverse transcription polymerase chain reaction (RT-PCR). Flow cytometry showed expression of some progenitor cell markers. In vivo study showed that the cells expressed both fetal and differentiated hepatocytes markers. Our study suggests new approaches to expand hepatic progenitor cells, analyze their fate in animal models aiming at cell therapy of hepatic diseases.

Cell-specific housekeeping role of lncRNAs in COVID-19-infected and recovered patients.

A plethora of studies have demonstrated the roles of lncRNAs in modulating disease severity and outcomes during infection. However, the spatio-temporal expression of these lncRNAs is poorly understood. In this study, we used single-cell RNA-seq to understand the spatio-temporal expression dynamics of lncRNAs across healthy, SARS-CoV-2-infected, and recovered individuals and their functional role in modulating the disease and recovery. We identified 203 differentially expressed lncRNAs, including cell type-specific ones like MALAT1, NEAT1, ZFAS1, SNHG7, SNHG8, and SNHG25 modulating immune function in classical monocyte, NK T, proliferating NK, plasmablast, naive, and activated B/T cells. Interestingly, we found invariant lncRNAs (no significant change in expression across conditions) regulating essential housekeeping functions (for example, HOTAIR, NRAV, SNHG27, SNHG28, and UCA1) in infected and recovered individuals. Despite similar repeat element abundance, variant lncRNAs displayed higher Alu content, suggesting increased interactions with proximal and distal genes, crucial for immune response modulation. The comparable repeat abundance but distinct expression levels of variant and invariant lncRNAs highlight the significance of investigating the regulatory mechanisms of invariant lncRNAs. Overall, this study offers new insights into the spatio-temporal expression patterns and functional roles of lncRNAs in SARS-CoV-2-infected and recovered individuals while highlighting the importance of invariant lncRNAs in the disease context.

Silver-doped hydroxyapatite laden chitosan-gelatin nanocomposite scaffolds for bone tissue engineering: an in-vitro and in-ovo evaluation.

Despite the advancements in bone tissue engineering, the majority of implant failures are caused due to microbial contamination. So, efforts are being made to develop biomaterial with antimicrobial property enhancing the regeneration of damaged bone tissue. In the present study, chitosan-gelatin (CG) scaffolds containing silver-doped hydroxyapatite (AgHAP) nanoparticles at 0.5%, 1.0% and 1.5% (w/v) were fabricated by lyophilization technique. The results confirmed the synthesis of AgHAP nanoparticles and showed interconnected porous structure of the nanocomposite scaffolds with 89%-75% porosity. Similarly, the swelling percentage, degradation behavior and compressive modulus of CG-AgHAP nanocomposite scaffolds were 1666%, 40% and 0.7 MPa, respectively. The developed nanocomposite scaffolds revealed better antimicrobial properties and bioactivity. The cell culture studies showed favorable viability of Wharton's jelly stem cells on CG-AgHAP nanocomposite scaffolds. CAM (chorioallantoic membrane) assay determined the angiogenic potential with better visualization of blood vessels in the CAM area. Hence, the obtained results confirmed that CG-AgHAP3 nanocomposite scaffold was the most suitable for bone tissue engineering applications among all scaffolds.

Dynamics of Whole Transcriptome Analysis (WTA) and Surface markers expression (AbSeq) in Immune Cells of COVID-19 Patients and Recovered captured through Single Cell Genomics.

Introduction: Single-cell multi-omics studies, such as multidimensional transcriptomics (whole transcriptomic analysis, WTA), and surface marker analysis (antibody sequencing, AbSeq), have turned out to be valuable techniques that offer inaccessible possibilities for single-cell profiling of mRNA, lncRNA, and proteins. Methods: We used this technique to understand the dynamics of mRNA and protein-level differences in healthy, COVID-19-infected and recovered individuals using peripheral blood mononuclear cells (PBMCs). Our results demonstrate that compared to mRNA expression, protein abundance is a better indicator of the disease state. Results: We demonstrate that compared to mRNA expression, protein abundance is a better indicator of the disease state. We observed high levels of cell identity and regulatory markers, CD3E, CD4, CD8A, CD5, CD7, GITR, and KLRB1 in healthy individuals, whereas markers related to cell activation, CD38, CD28, CD69, CD62L, CD14, and CD16 elevated in the SARS-CoV-2 infected patients at both WTA and AbSeq levels. Curiously, in recovered individuals, there was a high expression of cytokine and chemokine receptors (CCR5, CCR7, CCR4, CXCR3, and PTGRD2). We also observed variations in the expression of markers within cell populations under different states. Discussion: Furthermore, our study emphasizes the significance of employing an oligo-based method (AbSeq) that can help in diagnosis, prognosis, and protection from disease/s by identifying cell surface markers that are unique to different cell types or states. It also allows simultaneous study of a vast array of markers, surpassing the constraints of techniques like FACS to query the vast repertoire of proteins.

Characterization and angiogenic potential of CD146+ endometrial stem cells.

BACKGROUND: The human endometrium, lining the inner uterus, regenerates over 400 times uniquely during a woman's reproductive life. Endometrial stem cells (eSCs) enrich the tissue, resulting in a dense vascular network, significant angiogenic potential, and effective regeneration power. Being of natural angiogenic properties and proven effective in the treatment of vascular disorders, eSCs can be considered safe, reliable, and superior to other post-natal stem cells. Cluster of Differentiation 146 (CD146) has emerged as a pivotal marker associated with pericytes and endothelial cells for promoting angiogenesis. Endometrial cells with high CD146 expression could proliferate and differentiate into multiple lineages. This study will explore the role of CD146 in eSCs, focusing on the potential to boost the angiogenic and regenerative functions of the cells. The novelty of this study lies in the investigation of CD146 on eSC function, which may open new possibilities for eSC-based therapy in regenerative medicine and vascular disorders. METHODS: The study involved obtaining endometrial biopsies from active reproducing women to isolate and cultivate eSCs. eSCs were assessed for growth factor secretion pattern, characterized for their mesenchymal properties. Finally, eSCs were tested for their angiogenic potential by angiogenic gene expression profile and in-ovo chick embryo model. As aimed, to check the role of CD146 in eSC angiogenesis, CD146+ cells were magnetically sorted and cultured. The sorted cells underwent various analyses, including flowcytometry to identify mesenchymal markers and human growth factor panel to analyze growth factor secretion profiles The study evaluated the angiogenic potential of CD146 + cells using functional assays, including ring formation, endothelial differentiation, and wound scratch assays, to evaluate cell migration and healing capabilities. Molecular insights were obtained through chemokine and cytokine investigations In-ovo Chick model assay was conducted to check the angiogenic potential and evaluated through macroscopic as well as through immunohistochemistry. RESULT: Endometrial stem cells (eSCs) were successfully isolated using a combination of mechanical and enzymatic digestion, followed by culturing in complete DMEM media. The secretion profile of eSCs revealed significant production of various angiogenic growth factors, including Granulocyte macrophage colony-stimulating factor (GM-CSF), granulocyte colony stimulating factor (G-CSF), Vascular endothelial growth factor (VEGF), Fibroblast growth factors (FGF), and Platelet derived growth factor AA (PDGF-AA). The angiogenic gene profile indicated upregulation of several angiogenic genes in eSCs. The mesenchymal nature of eSCs was demonstrated through surface marker analysis (Cluster of differentiation 73, Cluster of differentiation 90, Cluster of differentiation 105) and trilineage differentiation. The in-ovo chick model confirmed the angiogenic potential of eSCs. CD146+ cells, isolated via magnetic sorting, exhibited enhanced angiogenic potential. These cells secreted significant levels of angiogenic growth factors such as VEGF. In Matrigel assays, CD146+ cells formed endothelial ring structures more rapidly and persistently than unsorted eSCs. Semi-quantitative PCR confirmed their endothelial differentiation. CD146+ cells express various angiogenic chemokines such as CXCL5, CXCL8, CCL3, and CCL20 and cytokines such as GM-CSF, Interleukin-1ß (IL-1ß), Interleukin-6 (IL-6), PDGF AA/BB, Epidermal growth factor (EGF), Endothelin 1, Angiopoietin. In-ovo chick model assay showed that CD146+ cells had superior angiogenesis, with more nodes, junctions, and segments compared to eSCs and controls. Immunohistochemistry confirmed increased expression of endothelial markers (Cluster of differentiation 31, VEGF, Vascular associated protein (VAP), Von Willebrand factor (vWF) in CD146+ cells. CONCLUSION: The study highlights the angiogenic potential of endometrial stem cells, particularly the CD146+ cell population. These cells promote angiogenesis, secreting growth factors and forming stable blood vessel structures. CD146+ cells have higher expression levels of VEGF and TGF-α, key factors in angiogenesis. This suggests CD146+ eSCs may be promising for therapeutic applications in vascular diseases requiring angiogenesis. Further research is needed.

A biologically functional bioink based on extracellular matrix derived collagen for 3D printing of skin.

Over the past few years, several advancements have been made to develop artificial skin that mimics human skin. Artificial skin manufactured using 3D printing technology that includes all epidermal and dermal components, such as collagen, may offer a viable solution. The skin-specific bioink was derived from digested chicken skin and incorporated into PVA (polyvinyl alcohol) and gelatin. The prepared bioink was further analyzed for its structure, stability, biocompatibility, and wound healing potential in in vitro, in ovo, and in vivo models. The 3D-printed skin showed excellent mechanical properties. In vitro scratch assays showed the proliferation and migration of cells within 24 h. In an in ovo assay, the 3D-printed skin facilitated the attachment of cells to the scaffolds. In the animal study, the quick cellular recruitment at the injury site accelerated wound healing. Further, hydroxyproline content was estimated to be 0.9-1.2 mg/ml, and collagen content was 7.5 %, which confirmed the epithelization. The relative expressions of MMP-9, COMP, TNF-α, and IL-6 genes were found to be increased compared to the control. These results demonstrate that 3D bioprinting represents a suitable technology to generate bioengineered skin for therapeutic and industrial applications in an automated manner.

Transplantation of Human Placenta Derived Mitochondria Promotes Cell Communication in Endometrium in a Murine Model of Disturbed Endometrium.

OBJECTIVES: Herein, we investigated the regenerative potential of functional mitochondria to restore endometrial injury. METHODS: The endometrium was disturbed with an intrauterine injection of 95% ethanol. Regeneration of the disturbed endometrium was achieved by transplantation of human placenta derived mitochondria followed by thrombin activated platelet rich plasma (hMTx). The transplantation method provided a biomimetic gel layer that stabilized and supported the functionality of the transplanted mitochondria to flourish regeneration of the disturbed endometrium. The presence of engrafted Rhodamine B labelled mitochondria was quantified at 12, 24, 48, and 72 h after transplantation. RESULTS: Detection of human-specific mitochondria mRNA in recipient rat uterus showed significant up-regulation of MT ATP-8, MT COX-1, MT COX -3, MT COX -2, MT ATP-6 (p = 0.009) in the hMTx treated group compared to the disturbed endometrium group. The hMTx group demonstrated showed regeneration through increased expressions of α-SMA, CK-18, CK-19, Connexin-40, E Cadherin, Claudin-1, Zona Occludin as compared with disturbed endometrium group. Experimental hMTx endometrial cells had significantly higher values of activities of NADH, NADPH, Cytochrome B5, Cytochrome P450, Complex I, Complex II, Complex III, Complex IV compared with disturbed endometrium indicating the regeneration of damaged endometrial cells at 72 h. CONCLUSIONS: Intrauterine hMTx was accounted to improve endometrial junction protein thus regeneration in the disturbed endometrium. Our Data provide the first evidence that hMTx promotes endometrial regeneration in the disturbed endometrium, paving the way for the development of a novel approach to human endometrial regeneration.

Dysregulated metal ion homeostasis underscores non-canonical function of CD8+ T cell during COVID-19.

Introduction: Several efforts have been made to describe the complexity of T cell heterogeneity during the COVID-19 disease; however, there remain gaps in our understanding in terms of the granularity within. Methods: For this attempt, we performed a single-cell transcriptomic analysis of 33 individuals (4 healthy, 16 COVID-19 positive patients, and 13 COVID-19 recovered individuals). Results: We found CD8+ T cell-biased lymphopenia in COVID-19 patients compared to healthy and recovered individuals. We also found an optimal Th1/Th2 ratio, indicating an effective immune response during COVID-19. Expansion of activated CD4+ T and NK T was detected in the COVID-19-positive individuals. Surprisingly, we found cellular and metal ion homeostasis pathways enriched in the COVID-19-positive individuals compared to the healthy and recovered in the CD8+ T cell populations (CD8+ TCM and CD8+ TEM) as well as activated CD4+ T cells. Discussion: In summary, the COVID-19-positive individuals exhibit a dynamic T cell mediated response. This response may have a possible association with the dysregulation of non-canonical pathways, including housekeeping functions in addition to the conventional antiviral immune response mediated by the T cell subpopulation. These findings considerably extend our insights into the heterogeneity of T cell response during and post-SARS-CoV-2 infection.

Single-cell RNA-Seq reveals intracellular microbial diversity within immune cells during SARS-CoV-2 infection and recovery.

Intracellular microorganisms, like viruses, bacteria, and fungi, pose challenges in detection due to their non-culturable forms. Transcriptomic analysis at cellular level enables exploration of distributions and the impact of these microorganisms on host cells, a domain that remains underexplored because of methodological limitations. Single-cell technology shows promise in addressing this by capturing polyadenine-tailed transcripts, because recent studies confirmed polyadenylation in microbial transcriptomes. We utilized single-cell RNA-seq from PBMCs to probe intracellular microbes in healthy, SARS-CoV-2-positive, and recovered individuals. Among 76 bacterial species detected, 16 showed significant abundance differences. Buchnera aphidicola, Streptomyces clavuligerus, and Ehrlichia canis emerged significantly in memory-B, Naïve-T, and Treg cells. Staphylococcus aureus, Mycoplasma mycoides, Leptospira interrogans, and others displayed elevated levels in SARS-CoV-2-positive patients, suggesting possible disease association. This highlights the strength of single-cell technology in revealing potential microorganism's cell-specific functions. Further research is essential for functional understanding of their cell-specific abundance across physiological states.

Decellularized small intestine scaffolds: a potential xenograft for restoration of intestinal perforation.

Small intestine perforation is a serious medical condition that requires immediate medical attention. The traditional course of treatment entails resection followed by anastomosis; however, it has complications such as small bowel syndrome (SBS), anastomotic leakage, and fistula formation. Here, a novel strategy is demonstrated, that utilizes the xenogeneic, decellularized goat small intestine as a patch for small intestine regeneration in cases of intestinal perforation. The goat small intestine scaffold underwent sodium dodecyl sulfate decellularization, which revealed consistent, quick, and effective decellularization. Decellularization contributed the least amount of extracellular matrix degradation while maintaining the intestinal architecture. By implanting the decellularized goat small intestine scaffolds (DGSIS) on the chorioallantoic membrane (CAM), no discernible loss of angiogenesis was seen in the CAM region, and this enabled the DGSIS to be evaluated for biocompatibility in ovo. The DGSIS was then xeno-transplanted as a patch on a small intestine perforation rat model. After 30 days post transplant, barium salt used as contrast gastrointestinal X-ray imaging revealed no leakage or obstruction in the small intestine. Histology, scanning electron microscopy, and immunohistochemistry assisted in analyzing the engraftment of host cells into the xeno patch. The xeno-patch expressed high levels of E-cadherin, α-smooth muscle actin (α-SMA), Occludin, Zonnula occluden (ZO-1), Ki 67, and Na+/K+-ATPase. The xeno-patch was consequently recellularized and incorporated into the host without causing an inflammatory reaction. As an outcome, decellularized goat small intestine was employed as a xenograft and could be suitable for regeneration of the perforated small intestine.

Cerium oxide nanoparticles disseminated chitosan gelatin scaffold for bone tissue engineering applications.

Cell-free and cell-loaded constructs are used to bridge the critical-sized bone defect. Oxidative stress at the site of the bone defects is a major interference that slows bone healing. Recently, there has been an increase in interest in enhancing the properties of three-dimensional scaffolds with free radical scavenging materials. Cerium oxide nanoparticles (CNPs) can scavenge free radicals due to their redox-modulating property. In this study, freeze-drying was used to fabricate CG-CNPs nanocomposite scaffolds using gelatin (G), chitosan (C), and cerium oxide nanoparticles. Physico-chemical, mechanical, and biological characterization of CG-CNPs scaffolds were studied. CG-CNPs scaffolds demonstrated better results in terms of physicochemical, mechanical, and biological properties as compared to CG-scaffold. CG-CNPs scaffolds were cyto-friendly to MC3T3-E1 cells studied by performing in-vitro and in-ovo studies. The scaffold's antimicrobial study revealed high inhibition zones against Gram-positive and Gram-negative bacteria. With 79 % porosity, 45.99 % weight loss, 178.25 kPa compressive modulus, and 1.83 Ca/P ratio, the CG-CNP2 scaffold displays the best characteristics. As a result, the CG-CNP2 scaffolds are highly biocompatible and could be applied to repair bone defects.

Surgical cotton microfibers loaded with proteins and apatite: A potential platform for bone tissue engineering.

Tissue engineering has emerged as the best alternative to replacing damaged tissue/organs. However, the cost of scaffold materials continues to be a significant obstacle; thus, developing inexpensive scaffolds is strongly encouraged. In this study, cellulose microfibers (C), gelatin (G), egg white (EW), and nanohydroxyapatite (nHA) were assembled into a quaternary scaffold using EDC-NHS crosslinking, followed by freeze-drying method. Cellulose microfibers as a scaffold have only received a limited amount of research due to the absence of an intrinsic three-dimensional structure. Gelatin, more likely to interact chemically with collagen, was used to provide a stable structure to the cellulose microfibers. EW was supposed to provide the scaffold with numerous cell attachment sites. nHA was chosen to enhance the scaffold's bone-bonding properties. Physico-chemical, mechanical, and biological characterization of scaffolds were studied. In-vitro using MG-63 cells and in-ovo studies revealed that all scaffolds were biocompatible. The results of the DPPH assay demonstrate the ability of CGEWnHA to reduce free radicals. The CGEWnHA scaffold exhibits the best properties with 56.84 ± 28.45 µm average pore size, 75 ± 1.4 % porosity, 39.23 % weight loss, 109.19 ± 0.98 kPa compressive modulus, and 1.72 Ca/P ratio. As a result, the constructed CGEWnHA scaffold appears to be a viable choice for BTE applications.