The aim of the present study was to examine whether combining strength and endurance training would promote better improvements in neuromuscular characteristics and functional abilities than endurance training alone in obese adolescent girls enrolled in a weight-reduction program. Twenty-four obese adolescent girls (12-15聽years) volunteered to participate in a 9-month training program. Participants were allocated into two groups following either (i) combined training (endurance聽+聽strength; E+ST) or (ii) endurance training (ET) program. Absolute and specific maximal torque, muscle size, and maximal voluntary activation level (VA) of the knee extensor (KE) and plantar flexor (PF) muscles were assessed. Moreover, functional abilities such as balance and fatigability during a maximal isometric intermittent contraction test of the KE muscles were measured before and after the intervention. The force of the adductor pollicis (AP) muscles was used as a control to account for any effect of growth or mechanical unloading on neuromuscular properties and muscle size. While absolute and specific torque of the KE (+14.7聽卤聽12.1% and +14.4聽卤聽15.5%; p聽<聽0.05) and PF (+19.2聽卤聽16.7% and +18.3聽卤聽17.5%; p聽<聽0.001) muscles increased in the E+ST group, PF torque decreased, and KE torque did not change in the ET group (-22.6聽卤聽10.5% and -15.0聽卤聽17.2%; p聽<聽0.001). Moreover, the VA of the KE muscles increased for the E+ST (+6.1聽卤聽5.6%; p聽<聽0.01) group and decreased for the ET group (-5.4聽卤聽5.4%; p聽<聽0.05). In contrast, VA remained similar in the PF muscles for both groups. The number of repetitions during the fatigability test increased in the ET group (38.4聽卤聽22.3 vs. 84.1聽卤聽33.3; p聽=聽0.032) and was unchanged in the E+ST group (50.8聽卤聽14.1 vs. 54.2聽卤聽37.8), but it was associated with a higher force level. Moreover, balance improved in the E+ST group, but not in the ET group. To conclude, physical training combining strength and endurance training promoted larger improvement in neuromuscular characteristics and functional abilities than endurance training alone in obese adolescent girls. Greater neuromuscular adaptations resulting from the E+ST training may be beneficial for preserving or even increasing functional abilities and possibly induce greater engagement in the active lifestyle of obese adolescents. However, the endurance component seems necessary in training programs to reduce fatigability during daily living activities.

Conditioned medium from cultured fibroblast cells is recognized to promote wound healing and growth through the secretion of enzymes, extracellular matrix proteins, and various growth factors and cytokines. The objective of this study was to profile the secreted proteins present in nasal fibroblast conditioned medium (NFCM). Nasal fibroblasts isolated from human nasal turbinates were cultured for 72聽h in Defined Keratinocytes Serum Free Medium (DKSFM) or serum-free F12: Dulbecco's Modified Eagle's Medium (DMEM) to collect conditioned medium, denoted as NFCM_DKSFM and NFCM_FD, respectively. SDS-PAGE was performed to detect the presence of protein bands, followed by MALDI-TOF and mass spectrometry analysis. SignalP, SecretomeP, and TMHMM were used to identify the secreted proteins in conditioned media. PANTHER Classification System was performed to categorize the protein according to protein class, whereas STRING 10 was carried out to evaluate the predicted proteins interactions. SDS-PAGE results showed the presence of various protein with molecular weight ranging from ~10聽kDa to ~260聽kDa. Four protein bands were identified using MALDI-TOF. The analyses identified 104, 83, and 7 secreted proteins in NFCM_FD, NFCM_DKSFM, and DKSFM, respectively. Four protein classes involved in wound healing were identified, namely calcium-binding proteins, cell adhesion molecules, extracellular matrix proteins, and signaling molecules. STRING10 protein prediction successfully identified various pathways regulated by secretory proteins in NFCM. In conclusion, this study successfully profiled the secreted proteins of nasal fibroblasts and these proteins are predicted to play important roles in RECs wound healing through various pathways.

INTRODUCTION: The lung is a complex organ composed of numerous cell types. Exposure to air pollutants, cigarette smoke, bacteria, viruses, and many others may cause injury to the epithelial cells that line the conducting airways and alveoli. Organoids are the 3D self-organising structures grown from stem cells and generated from adult stem and progenitor cells. Lung organoids are fascinating tools to investigate human lung development in vitro. The objective of this study was to establish a rapid method for generating lung organoids with a direct culture strategy. METHODS: Trachea and lung organoids were derived from mixed cell populations of mice primary airway epithelial cells, fibroblasts, and lung microvascular endothelial cells and directly digested from the whole cell population in the distal lung. RESULTS: The formation of spheres appeared as early as 3聽days and continued to proliferate until day 5. The generation of trachea and lung organoids self-organised into discrete epithelial structures was formed within less than 10聽days. CONCLUSION: We conclude that researchers will be able to examine cellular involvement during organ formation and molecular networks because organoids come in a variety of morphologies and stages of development, and this organoid protocol may be used for modelling lung diseases as a platform for therapeutic purposes and suitable for personalised medicine for respiratory diseases.

INTRODUCTION: Recent studies have demonstrated that adipose tissue-derived stem cell (ADSC) transplantation could promote neoangiogenesis in various ischemic diseases. However, as whole cells, ADSCs have some disadvantages, such as shipping and storage issues, high costs, and controversies related to the fates of grafted cells in the recipients. Therefore, this study aimed to investigate the effects of intravenously infused exosomes purified from human ADSCs on ischemic disease in a murine hindlimb ischemia model. METHODS: ADSCs were cultured in exosome-free medium for 48聽h before the conditioned medium was collected for exosome isolation by ultracentrifugation. The murine ischemic hindlimb models were created by cutting and burning the hindlimb arteries. Exosomes were intravenously infused into murine models (ADSC-Exo group), with phosphate-buffered saline (PBS) used as a placebo (PBS group). Treatment efficacy was determined using a murine mobility assay (frequency of pedaling in water per 10聽s), peripheral blood oxygen saturation (SpO2 index), and the recovery of vascular circulation by trypan blue staining. The formation of blood vessels was shown by X-ray. Expression levels of genes related to angiogenesis and muscle tissue repair were quantified by quantitative reverse-transcription polymerase chain reaction. Finally, H&E staining was used to determine the histological structure of muscle in the treatment and placebo groups. RESULTS: The rates of acute limb ischemia in the PBS and ADSC-Exo injection groups were 66% (9/16 mice) and 43% (6/14 mice), respectively. The mobility of the limbs 28聽days after surgery was significantly different between the ADSC-Exo treatment group (41聽卤聽1 times/10聽s) and the PBS group (24聽卤聽1 times/10聽s; n聽=聽3; p聽<聽0.05). Peripheral blood oxygen saturation 21聽days after treatment was 83.83%聽卤聽2.02% in the PBS group and 83%聽卤聽1.73% in the ADSC-Exo treatment group, and the difference was not statistically significant (n聽=聽3, p聽>聽0.05). On day 7 after treatment, the time required to stain the toes after trypan blue injection was 20.67聽卤聽12.5聽s and 85聽卤聽7.09聽s in the ADSC-Exo and PBS groups, respectively (n聽=聽3, p聽<聽0.05). On day 3 after the operation, the expression of genes promoting angiogenesis and muscle remodeling, such as Flk1, Vwf, Ang1, Tgfb1, Myod, and Myf5, was increased 4-8 times in the ADSC-Exo group compared with the PBS group. No mice in either group died during the experimental period. CONCLUSIONS: These results revealed that intravenous infusion of human ADSC-derived exosomes is a safe and effective method to treat ischemic disease, especially hindlimb ischemia, by promoting angiogenesis and muscle regeneration.

INTRODUCTION: The pro-inflammatory cytokine interferon-gamma (IFN-纬) is reported to be an agent that boosts the immune modulation of mesenchymal stem cells (MSCs). However, the effects of IFN-纬 on the chondrogenic potential of treated MSCs have not been evaluated in depth. This study aimed to evaluate the effects of IFN-纬 on the immune modulation and chondrogenic potential of human umbilical cord-derived MSCs (hUC-MSCs). METHODS: UC-MSCs were isolated and expanded following published protocols. They were characterized as MSCs before their use in further experiments. The UC-MSCs were treated with IFN-纬 at 10聽ng/mL for 48聽h. Changes in phenotype were investigated based on changes in MSC markers, immunomodulatory genes (TGF-尾, IL-4, and IDO) for immune modulation, and cartilage-related genes during the induction of differentiation (Col1a2, Col2a1, Sox9, Runx2, and Acan) for chondrogenic potential. RESULTS: IFN-纬-treated UC-MSCs maintained MSC markers and exhibited decreased expression of transcriptional regulatory factors in chondrogenesis (Sox9 and Runx2) and the extracellular matrix-specific genes Col1a2 and Acan but not Col2a1 compared to non-treated cells (p聽<聽0.05). Furthermore, the immunomodulatory capability of IFN-纬-treated UC-MSCs was clearly revealed through their increased expression of IDO and IL-4 and decreased expression of TGF-尾 compared to non-treated cells (p聽<聽0.05). CONCLUSION: This study demonstrated that UC-MSCs treated with IFN-纬 at 10聽ng/mL had reduced expression of chondrocyte-specific genes; however, they maintained multi-lineage differentiation and exhibited immunomodulatory properties.

The thymus is the main immune organ in the body. However, the thymus gradually degenerates in early life, leading to a reduction in T-cell production and a decrease in immune function. Mesenchymal stem cells (MSCs) are a promising alternative for the treatment of thymus senescence due to their homing ability to the site of inflammation and their paracrine, anti-inflammatory, and antioxidant properties. However, the heterogeneity, difficulty of survival in vivo, short residence time, and low homing efficiency of the injected MSCs affect the clinical therapeutic effect. This article reviews strategies to improve the efficacy of mesenchymal stem cell therapy, including the selection of appropriate cell doses, transplantation frequency, and interval cycles. The survival rate of MSCs can be improved to some extent by improving the infusion mode of MSCs, such as simulating the in vivo environment, applying the biological technology of hydrogels and microgels, and iron oxide labeling technology, which can improve the curative effect and homing of MSCs, promote the regeneration of thymic epithelial cells, and restore the function of the thymus.

Apoptotic and healthy cells of domestic animals release membrane-enclosed particles from their plasma membrane. These special structures, called extracellular vesicles, play an important role in intercellular communication. In the past, it was believed that their function was mainly to dispose unwanted cell contents and to help maintain cell homeostasis. However, we now know that they have important roles in health and disease and have diagnostic value as well as great potential for therapy in veterinary medicine. Extracellular vesicles facilitate cellular exchanges by delivering functional cargo molecules to nearby or distant tissues. They are produced by various cell types and are found in all body fluids. Their cargo reflects the state of the releasing parent cell, and despite their small size, this cargo is extraordinarily complex. Numerous different types of molecules contained in vesicles make them an extremely promising tool in the field of regenerative veterinary medicine. To further increase research interest and discover their full potential, some of the basic biological mechanisms behind their function need to be better understood. Only then will we be able to maximize the clinical relevance for targeted diagnostic and therapeutic purposes in various domestic animal species.

Hepatocellular carcinoma (HCC) is a primary liver malignancy that accounts for the majority of liver cancer cases, with multiple risk factors including chronic hepatitis B and C infections, alcohol abuse, and non-alcoholic fatty liver disease (NAFLD). Despite advancements in diagnosis and treatment, the survival rate of patients with advanced HCC remains low, creating an urgent need for new therapeutic targets and strategies.One biological process crucial to HCC progression is the epithelial-mesenchymal transition (EMT). EMT is a process that enables epithelial cells to acquire mesenchymal properties, including motility and invasiveness, by losing their cell-cell adhesion. Various signaling pathways, including TGF-尾, Wnt/尾-catenin, and Notch, have been implicated in regulating EMT in HCC.To inhibit EMT, targeted therapeutic approaches have been developed, and preclinical studies suggest that the inhibition of the TGF-尾, Wnt/尾-catenin, and Notch signaling pathways is promising. TGF-尾 receptor inhibitors, Wnt/尾-catenin pathway inhibitors, and gamma-secretase inhibitors have shown efficacy in preclinical studies by inhibiting EMT and reducing tumor growth in HCC models. However, further clinical studies are necessary to determine their effectiveness in human patients.In addition to these approaches, further research is needed to identify other novel therapeutic targets and develop new treatment strategies for HCC. This review emphasizes the critical role of EMT in HCC progression and highlights the potential of targeting the TGF-尾, Wnt/尾-catenin, and Notch signaling pathways to inhibit EMT and reduce tumor growth in HCC. Future studies and clinical trials are necessary to validate these therapeutic strategies and develop effective treatments for HCC.

The regulation of glucose homeostasis and insulin secretion by pancreatic 尾-cells, when disturbed, will result in diabetes mellitus. Replacement of dysfunctional or lost 尾-cells with fully functional ones can tackle the problem of 尾-cell generation in diabetes mellitus. Various pancreatic-specific genes are expressed during different stages of development, which have essential roles in pancreatogenesis and 尾-cell formation. These factors play a critical role in cellular-based studies like transdifferentiation or de-differentiation of somatic cells to multipotent or pluripotent stem cells and their differentiation into functional 尾-cells. This work gives an overview of crucial transcription factors expressed during various stages of pancreas development and their role in 尾-cell specification. In addition, it also provides a perspective on the underlying molecular mechanisms.

Cancer stem cells (CSC) have unique characteristics which include self-renewal, multi-directional differentiation capacity, quiescence/dormancy, and tumor-forming capability. These characteristics are referred to as the "stemness" properties. Tumor microenvironment contributes to CSC survival, function, and remaining them in an undifferentiated state. CSCs can form malignant tumors with heterogeneous phenotypes mediated by the tumor microenvironment. Therefore, the crosstalk between CSCs and tumor microenvironment can modulate tumor heterogeneity. CSCs play a crucial role in several biological processes, epithelial-mesenchymal transition (EMT), autophagy, and cellular stress response. In this chapter, we focused characteristics of cancer stem cells, reprogramming strategies cells into CSCs, and then we highlighted the contribution of CSCs to therapy resistance and cancer relapse and their potential of therapeutic targeting of CSCs.

Advances in cellular reprogramming articulated the path for direct cardiac lineage conversion, bypassing the pluripotent state. Direct cardiac reprogramming attracts major attention because of the low or nil regenerative ability of cardiomyocytes, resulting in permanent cell loss in various heart diseases. In the field of cardiology, balancing this loss of cardiomyocytes was highly challenging, even in the modern medical world. Soon after the discovery of cell reprogramming, direct cardiac reprogramming also became a promising alternative for heart regeneration. This review mainly focused on the various direct cardiac reprogramming approaches (integrative and non-integrative) for the derivation of induced autologous cardiomyocytes. It also explains the advancements in cardiac reprogramming over the decade with the pros and cons of each approach. Further, the review highlights the importance of clinically relevant (non-integrative) approaches and their challenges for the prospective applications for personalized medicine. Apart from direct cardiac reprogramming, it also discusses the other strategies for generating cardiomyocytes from different sources. The understanding of these strategies could pave the way for the efficient generation of integration-free functional autologous cardiomyocytes through direct cardiac reprogramming for various biomedical applications.

The plasma membrane is not a uniform phospholipid bilayer; it has specialized membrane nano- or microdomains called lipid rafts. Lipid rafts are small cholesterol and sphingolipid-rich plasma membrane islands. Although their existence was long debated, their presence in the plasma membrane of living cells is now well accepted with the advent of super-resolution imaging techniques. It is interesting to note that lipid rafts function to compartmentalize receptors and their regulators and substantially modulate cellular signaling. In this review, we will examine the role of lipid rafts and caveolae-lipid raft-like microdomains with a distinct 3D morphology-in cellular signaling. Moreover, we will investigate how raft compartmentalized signaling regulates diverse physiological processes such as proliferation, apoptosis, immune signaling, and development. Also, the deregulation of lipid raft-mediated signaling during tumorigenesis and metastasis will be explored.

Metal ions can be both essential components of cells as well as potential toxins if present in excess. Organisms utilize a variety of protein systems to maintain the concentration of metal ions within the appropriate range for cellular function, and to avoid concentrations where cellular damage can occur. In bacteria, numerous proteins contribute to copper homeostasis, including copper transporters, chelators, and redox enzymes. The genes that encode these proteins are often found in clusters, thus providing modular components that work together to achieve homeostasis. A better understanding of how these components function and cooperate to achieve metal ion resistance is needed, given the extensive use of metal ions, including copper, as broad-spectrum biocides in a variety of clinical and environmental settings. The copG gene is a common component of such copper resistance clusters, but its contribution to copper resistance is not well understood. In this review the available information about the CopG protein encoded by this gene is summarized. Comparison of the recent structure to diverse copper-containing metallochaperones, metalloenzymes, and electron transfer proteins suggests that CopG is a redox enzyme that uses multiple copper ions as active site redox cofactors to act on additional copper ion substrates. Mechanisms for both oxidase and reductase activity are proposed, and the biological advantages that these activities can contribute in conjunction with existing systems are described.

Stem cells, which can self-renew and differentiate into different cell types, have become the keystone of regenerative medicine due to these properties. With the achievement of superior clinical results in the therapeutic approaches of different diseases, the applications of these cells in the treatment of genetic diseases have also come to the fore. Foremost, conventional approaches of stem cells to genetic diseases are the first approaches in this manner, and they have brought safety issues due to immune reactions caused by allogeneic transplantation. To eliminate these safety issues and phenotypic abnormalities caused by genetic defects, firstly, basic genetic engineering practices such as vectors or RNA modulators were combined with stem cell-based therapeutic approaches. However, due to challenges such as immune reactions and inability to target cells effectively in these applications, advanced molecular methods have been adopted in ZFN, TALEN, and CRISPR/Cas genome editing nucleases, which allow modular designs in stem cell-based genetic diseases' therapeutic approaches. Current studies in genetic diseases are in the direction of creating permanent treatment regimens by genomic manipulation of stem cells with differentiation potential through genome editing tools. In this chapter, the stem cell-based therapeutic approaches of various vital genetic diseases were addressed wide range from conventional applications to genome editing tools.

Chronic obstructive pulmonary disease (COPD) patients manifest muscle dysfunction and impaired muscle oxidative capacity, which result in reduced exercise capacity and poor health status. The aim of this study was to compare the physical performance, systemic inflammation, and oxidative stress of patients with moderate COPD, and to associate physical performance with inflammatory and oxidative stress plasma markers. Twenty CONTROL (n聽=聽10) and moderate COPD (n聽=聽10) patients participated in this study. Systematic inflammation and oxidative stress plasma markers, maximal aerobic capacity (VO2peak), and maximal isometric strength (MVIC) of the knee extensor (KE) muscles were measured. VO2peak was 31.3% greater in CONTROL compared to COPD (P聽=聽0.006). The MVIC strength of the KE was 43.9% greater in CONTROL compared to COPD (P聽=聽0.002). Tumor necrosis factor-alpha (TNF-伪) was 79.6% greater in COPD compared to CONTROL (P聽<聽0.001). Glutathione peroxidase activity (GPx) activity was 27.5% lesser in COPD compared to CONTROL (P聽=聽0.05). TNF-伪 concentration was correlated with KE MVC strength (R聽=聽-0.48; P聽=聽0.045) and VO2peak (R聽=聽-0.58; P聽=聽0.01). Meanwhile, malondialdehyde (MDA) and GPx activity were not associated with KE strength or VO2peak (P聽=聽0.74 and P聽=聽0.14, respectively). COPD patients showed lesser muscle strength and aerobic capacity than healthy control individuals. Furthermore, patients with COPD showed greater systemic inflammation and lesser antioxidant capacity than healthy counterparts. A moderate association was evident between levels of systemic inflammation and physical performance variables.

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, 伪-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.

Environmental nutrients control bacterial biofilm homeostasis, by regulating the intracellular levels of c-di-GMP. One component transducers can sense different classes of small molecules through a periplasmic domain; the nutrient recognition triggers the subsequent regulation of the downstream cytosolic diguanylate cyclase (GGDEF) or phosphodiesterase (EAL) domains, via transmembrane helix(ces), to finally change c-di-GMP levels.Protein studies on such transducers have been mainly carried out on isolated domains due to the presence of the transmembrane portion. Nevertheless, the cleavage of GGDEF and EAL-containing proteins could be detrimental since both tertiary and quaternary structures could be allosterically controlled; to by-pass this limitation, studies on the corresponding full-length proteins are highly desired.We have in silico selected a GGDEF-EAL transducer from Dyella thiooxydans (ann. A0A160N0B7), whose periplasmic binding domain was predicted to bind to arginine, a nutrient often associated with chronic infections and biofilm. This protein has been used as an in vitro tool for the identification of the best approach for its isolation, including (i) protein engineering to produce a water-soluble version via QTY (Glutamine, Threonine, and Tyrosine) code or (ii) nanodiscs assembly. The results on this "prototype" may represent the proof-of-concept for future isolation of other transmembrane proteins sharing the same architecture, including more complex nutrient-based transducers controlling c-di-GMP levels.

This study investigates the in vitro antibacterial activity of coconut oil on selected clinical and pure bacterial isolates. Clinical samples were isolated from the people of Ras Al Khaimah, United Arab Emirates. Biochemical examination of the microorganisms was done according to standard methods. Pure bacterial cultures were provided from LTA srl Italia. In this research work, an effort has been made to highlight the valuable properties of Cocos nucifera oil, in order to rationalize the use of coconut oil against bacteria. Experiments were performed by agar well diffusion method. Ciprofloxacin was used as a standard antibiotic. The assay of antibacterial activity of clinical isolate of Streptococcus species showed the highest susceptibility to coconut oil while Escherichia coli had the least. This study endorses the use of coconut oil as therapeutic agent since it contains lauric acid which is bactericidal. The utilization of coconut oil should be promoted as a functional food and the use of coconut seed flesh in our diets should be encouraged for health-supporting functions. Further studies should be done on the oil and its derivatives both in vitro and in vivo to unveil their mechanism of action.

INTRODUCTION: Exosomes derived from mesenchymal stem cells (MSCs) are crucial mediators of the paracrine effects as well as tissue repair and have promising clinical applications. They enhance tissue regeneration by reducing inflammatory responses, enhancing proliferation, inhibiting apoptosis, and stimulating angiogenesis. This study aimed to evaluate the mechanism of angiogenesis supported by exosomes derived from MSCs. METHODS: Exosomes were isolated via ultracentrifugation of a conditioned medium collected from human umbilical cord MSC (hUCMSC) cultures. These exosomes were characterized using transmission electron microscopy, and the expression of specific markers (CD9, CD81, and CD63) was evaluated. To understand the mechanism of angiogenesis, we evaluated the effects of exosomes in endothelial cells (HUVECs). The obtained exosomes were supplemented at a dose of 20聽渭g/mL into two kinds of culture media for HUVECs (M200 medium and endothelial cell growth medium), while phosphate-buffered saline was added to these media as a control. The effects of the exosomes were evaluated based on the formation of a tubular structure in the culture and the expression of angiogenic genes (MMP-2, Ephrin B2, Ephrin B4, Flk1, Flt1, VWF, VE-cadherin, CD31, ANG1, ANG2, and HGF) via RT-PCR. RESULTS: The exosomes were obtained from the hUCMSCs at a concentration of 0.7聽卤聽0.029聽渭g/mL. They accelerated the formation of new blood vessels by upregulating HGF, VWF, CD31, Flt1, and Flk1 (especially VWF and Flt1). CONCLUSION: Exosomes derived from hUCMSCs can promote angiogenesis through upregulation of VWF and Flt1 in endothelial cells.

Since insulin deficiency results from pancreatic beta-cell destruction, all type 1 and most type 2 diabetes patients eventually require life-long insulin injections. Insulin gene synthesis could also be impaired due to insulin gene mutations as observed in diabetic patients with MODY 10. At this point, insulin gene therapy could be very effective to recompense insulin deficiency under these circumstances. For this reason, an HIV-based lentiviral vector carrying the insulin gene under the control of insulin promoter (LentiINS) was generated, and its therapeutic efficacy was tested in a beta-cell transplant model lacking insulin produced by CRISPR/Cas9-mediated genetically engineered pancreatic beta cells. To generate an insulin knockout beta-cell transplant animal model of diabetes, a dual gene knockout plasmid system involving CRISPR/Cas9 was transfected into a mouse pancreatic beta cell line (Min6). Fluorescence microscopy and antibiotic selection were utilized to select the insulin gene knockout clones. Transplantation of the genetically engineered pancreatic beta cells under the kidney capsule of STZ-induced diabetic rats revealed LentiINS- but not LentiLacZ-infected Ins2KO cells transiently reduced hyperglycemia similar to that of MIN6 in diabetic animals. These results suggest LentiINS has the potential to functionally restore insulin production in an insulin knockout beta-cell transplant animal model of diabetes.