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1.
Engineered Zinc Finger Protein Targeting 2LTR Inhibits HIV Integration in Hematopoietic Stem and Progenitor Cell-Derived Macrophages: In Vitro Study.
Chupradit, K, Khamaikawin, W, Sakkhachornphop, S, Puaninta, C, Torbett, BE, Borwornpinyo, S, Hongeng, S, Wattanapanitch, M, Tayapiwatana, C
International journal of molecular sciences. 2022;(4)
Abstract
Human hematopoietic stem/progenitor cell (HSPC)-based gene therapy is a promising direction for curing HIV-1-infected individuals. The zinc finger protein (2LTRZFP) designed to target the 2-LTR-circle junction of HIV-1 cDNA was previously reported as an intracellular antiviral molecular scaffold that prevents HIV integration. Here, we elucidate the efficacy and safety of using 2LTRZFP in human CD34+ HSPCs. We transduced 2LTRZFP which has the mCherry tag (2LTRZFPmCherry) into human CD34+ HSPCs using a lentiviral vector. The 2LTRZFPmCherry-transduced HSPCs were subsequently differentiated into macrophages. The expression levels of pro-apoptotic proteins of the 2LTRZFPmCherry-transduced HSPCs showed no significant difference from those of the non-transduced control. Furthermore, the 2LTRZFPmCherry-transduced HSPCs were successfully differentiated into mature macrophages, which had normal phagocytic function. The cytokine secretion assay demonstrated that 2LTRZFPmCherry-transduced CD34+ derived macrophages promoted the polarization towards classically activated (M1) subtypes. More importantly, the 2LTRZFPmCherry transduced cells significantly exhibited resistance to HIV-1 integration in vitro. Our findings demonstrate that the 2LTRZFPmCherry-transduced macrophages were found to be functionally and phenotypically normal, with no adverse effects of the anti-HIV-1 scaffold. Our data suggest that the anti-HIV-1 integrase scaffold is a promising antiviral molecule that could be applied to human CD34+ HSPC-based gene therapy for AIDS patients.
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2.
Adipose tissue-derived stromal/stem cells + cholecalciferol: a pilot study in recent-onset type 1 diabetes patients.
Dantas, JR, Araújo, DB, Silva, KR, Souto, DL, de Fátima Carvalho Pereira, M, Luiz, RR, Dos Santos Mantuano, M, Claudio-da-Silva, C, Gabbay, MAL, Dib, SA, et al
Archives of endocrinology and metabolism. 2021;(3):342-351
Abstract
OBJECTIVE Adipose tissue-derived stromal/stem cells (ASCs) and vitamin D have immunomodulatory actions that could be useful for type 1 diabetes (T1D). We aimed in this study to investigate the safety and efficacy of ASCs + daily cholecalciferol (VIT D) for 6 months in patients with recent-onset T1D. METHODS In this prospective, dual-center, open trial, patients with recent onset T1D received one dose of allogenic ASC (1 × 106 cells/kg) and cholecalciferol 2,000 UI/day for 6 months (group 1). They were compared to patients who received chol-ecalciferol (group 2) and standard treatment (group 3). Adverse events were recorded; C-peptide (CP), insulin dose and HbA1c were measured at baseline (T0), after 3 (T3) and 6 months (T6). RESULTS In group 1 (n = 7), adverse events included transient headache (all), mild local reactions (all), tachycardia (n = 4), abdominal cramps (n = 1), thrombophlebitis (n = 4), scotomas (n = 2), and central retinal vein occlusion at T3 (n = 1, resolution at T6). Group 1 had an increase in basal CP (p = 0.018; mean: 40.41+/-40.79 %), without changes in stimulated CP after mixed meal (p = 0.62), from T0 to T6. Basal CP remained stable in groups 2 and 3 (p = 0.58 and p = 0.116, respectively). Group 1 had small insulin requirements (0.31+/- 0.26 UI/kg) without changes at T6 (p = 0.44) and HbA1c decline (p = 0.01). At T6, all patients (100%; n = 7) in group 1 were in honeymoon vs 75% (n = 3/4) and 50% (n = 3/6) in groups 2 and 3, p = 0.01. CONCLUSION Allogenic ASC + VIT D without immunosuppression was safe and might have a role in the preservation of β-cells in patients with recent-onset T1D. ClinicalTrials.gov: NCT03920397.
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3.
Protocol and Baseline Data on Renal Autologous Cell Therapy Injection in Adults with Chronic Kidney Disease Secondary to Congenital Anomalies of the Kidney and Urinary Tract.
Stavas, J, Diaz-Gonzalez de Ferris, M, Johns, A, Jain, D, Bertram, T
Blood purification. 2021;(4-5):678-683
Abstract
BACKGROUND Advanced cell therapies with autologous, homologous cells show promise to affect reparative and restorative changes in the chronic kidney disease (CKD) nephron. We present our protocol and preliminary analysis of an IRB-approved, phase I single-group, open-label trial that tests the safety and efficacy of Renal Autologous Cell Therapy (REACT; NCT04115345) in adults with congenital anomalies of the kidney and urinary tract (CAKUT). METHODS Adults with surgically corrected CAKUT and CKD stages 3 and 4 signed an informed consent and served as their "own" baseline control. REACT is an active biological ingredient acquired from a percutaneous tissue acquisition from the patient's kidney cortex. The specimen undergoes a GMP-compliant manufacturing process that harvests the selected renal cells composed of progenitors for renal repair, followed by image-guided locoregional reinjection into the patient's renal cortex. Participants receive 2 doses at 6-month intervals. Primary outcomes are stable renal function and stable/improved quality of life. Additional exploratory endpoints include the impact of REACT on blood pressure, vitamin D levels, hemoglobin, hematocrit and kidney volume by MRI analysis. RESULTS Four men and 1 woman were enrolled and underwent 5 cell injections. Their characteristics were as follows: mean 52.8 years (SD 17.7 years), 1 Hispanic, 4 non-Hispanic, and 5 white. There were no renal tissue acquisition, cell injection, or cell product-related complications at baseline. CONCLUSION REACT is demonstrating feasibility and patient safety in preliminary analysis. Autologous cell therapy treatment has the potential to stabilize or improve renal function in CAKUT-associated CKD to delay or avert dialysis. Patient enrollment and follow-up are underway.
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4.
Natural Compounds and PCL Nanofibers: A Novel Tool to Counteract Stem Cell Senescence.
Bellu, E, Cruciani, S, Garroni, G, Balzano, F, Satta, R, Montesu, MA, Fadda, A, Mulas, M, Sarais, G, Bandiera, P, et al
Cells. 2021;(6)
Abstract
Tissue homeostasis mainly depends on the activity of stem cells to replace damaged elements and restore tissue functions. Within this context, mesenchymal stem cells and fibroblasts are essential for maintaining tissue homeostasis in skin, in particular in the dermis. Modifications in collagen fibers are able to affect stem cell features. Skin properties can be significantly reduced after injuries or with aging, and stem cell niches, mainly comprising extracellular matrix (ECM), may be compromised. To this end, specific molecules can be administrated to prevent the aging process induced by UV exposure in the attempt to maintain a youngness phenotype. NanoPCL-M is a novel nanodevice able to control delivery of Mediterranean plant myrtle (Myrtus communis L.) extracts. In particular, we previously described that myrtle extracts, rich in bioactive molecules and nutraceuticals, were able to counteract senescence in adipose derived stem cells. In this study, we analyzed the effect of NanoPCL-M on skin stem cells (SSCs) and dermal fibroblasts in a dynamic cell culture model in order to prevent the effects of UV-induced senescence on proliferation and collagen depot. The BrdU assay results highlight the significantly positive effect of NanoPCL-M on the proliferation of both fibroblasts and SSCs. Our results demonstrate that-M is able to preserve SSCs features and collagen depot after UV-induced senescence, suggesting their capability to retain a young phenotype.
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5.
Human Granulosa Cells-Stemness Properties, Molecular Cross-Talk and Follicular Angiogenesis.
Dompe, C, Kulus, M, Stefańska, K, Kranc, W, Chermuła, B, Bryl, R, Pieńkowski, W, Nawrocki, MJ, Petitte, JN, Stelmach, B, et al
Cells. 2021;(6)
Abstract
The ovarian follicle is the basic functional unit of the ovary, comprising theca cells and granulosa cells (GCs). Two different types of GCs, mural GCs and cumulus cells (CCs), serve different functions during folliculogenesis. Mural GCs produce oestrogen during the follicular phase and progesterone after ovulation, while CCs surround the oocyte tightly and form the cumulus oophurus and corona radiata inner cell layer. CCs are also engaged in bi-directional metabolite exchange with the oocyte, as they form gap-junctions, which are crucial for both the oocyte's proper maturation and GC proliferation. However, the function of both GCs and CCs is dependent on proper follicular angiogenesis. Aside from participating in complex molecular interplay with the oocyte, the ovarian follicular cells exhibit stem-like properties, characteristic of mesenchymal stem cells (MSCs). Both GCs and CCs remain under the influence of various miRNAs, and some of them may contribute to polycystic ovary syndrome (PCOS) or premature ovarian insufficiency (POI) occurrence. Considering increasing female fertility problems worldwide, it is of interest to develop new strategies enhancing assisted reproductive techniques. Therefore, it is important to carefully consider GCs as ovarian stem cells in terms of the cellular features and molecular pathways involved in their development and interactions as well as outline their possible application in translational medicine.
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6.
In Vitro Osteoinductivity Assay of Hydroxylapatite Scaffolds, Obtained with Biomorphic Transformation Processes, Assessed Using Human Adipose Stem Cell Cultures.
Iaquinta, MR, Torreggiani, E, Mazziotta, C, Ruffini, A, Sprio, S, Tampieri, A, Tognon, M, Martini, F, Mazzoni, E
International journal of molecular sciences. 2021;(13)
Abstract
In this study, the in vitro biocompatibility and osteoinductive ability of a recently developed biomorphic hydroxylapatite ceramic scaffold (B-HA) derived from transformation of wood structures were analyzed using human adipose stem cells (hASCs). Cell viability and metabolic activity were evaluated in hASCs, parental cells and in recombinant genetically engineered hASC-eGFP cells expressing the green fluorescence protein. B-HA osteoinductivity properties, such as differentially expressed genes (DEG) involved in the skeletal development pathway, osteocalcin (OCN) protein expression and mineral matrix deposition in hASCs, were evaluated. In vitro induction of osteoblastic genes, such as Alkaline phosphatase (ALPL), Bone gamma-carboxyglutamate (gla) protein (BGLAP), SMAD family member 3 (SMAD3), Sp7 transcription factor (SP7) and Transforming growth factor, beta 3 (TGFB3) and Tumor necrosis factor (ligand) superfamily, member 11 (TNFSF11)/Receptor activator of NF-κB (RANK) ligand (RANKL), involved in osteoclast differentiation, was undertaken in cells grown on B-HA. Chondrogenic transcription factor SRY (sex determining region Y)-box 9 (SOX9), tested up-regulated in hASCs grown on the B-HA scaffold. Gene expression enhancement in the skeletal development pathway was detected in hASCs using B-HA compared to sintered hydroxylapatite (S-HA). OCN protein expression and calcium deposition were increased in hASCs grown on B-HA in comparison with the control. This study demonstrates the biocompatibility of the novel biomorphic B-HA scaffold and its potential use in osteogenic differentiation for hASCs. Our data highlight the relevance of B-HA for bone regeneration purposes.
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7.
Possible role of intestinal stem cells in the pathophysiology of irritable bowel syndrome.
El-Salhy, M
World journal of gastroenterology. 2020;(13):1427-1438
Abstract
The pathophysiology of irritable bowel syndrome (IBS) is not completely understood. However, several factors are known to play a role in pathophysiology of IBS such as genetics, diet, gut microbiota, gut endocrine cells, stress and low-grade inflammation. Understanding the pathophysiology of IBS may open the way for new treatment approaches. Low density of intestinal stem cells and low differentiation toward enteroendocrine cells has been reported recently in patients with IBS. These abnormalities are believed to be the cause of the low density of enteroendocrine cells seen in patients with IBS. Enteroendocrine cells regulate gastrointestinal motility, secretion, absorption and visceral sensitivity. Gastrointestinal dysmotility, abnormal absorption/secretion and visceral hypersensitivity are all seen in patients with IBS and haven been attributed to the low density the intestinal enteroendocrine cells in these patients. The present review conducted a literature search in Medline (PubMed) covering the last ten years until November 2019, where articles in English were included. Articles about the intestinal stem cells and their possible role in the pathophysiology of IBS are discussed in the present review. The present review discusses the assumption that intestinal stem cells play a central role in the pathophysiology of IBS and that the other factors known to contribute to the pathophysiology of IBS such as genetics, diet gut microbiota, stress, and low-grade inflammation exert their effects through affecting the intestinal stem cells. It reports further the data that support this assumption on genetics, diet, gut microbiota, stress with depletion of glutamine, and inflammation.
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8.
Simulated microgravity significantly altered metabolism in epidermal stem cells.
Li, BB, Chen, ZY, Jiang, N, Guo, S, Yang, JQ, Chai, SB, Yan, HF, Sun, PM, Hu, G, Zhang, T, et al
In vitro cellular & developmental biology. Animal. 2020;(3):200-212
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Abstract
Simulated microgravity can significantly affect various cell types and multiple systems of the human body, such as cardiovascular system, skeletal muscle system, and immune system, and is known to cause anemia and loss of electrolyte and fluids. Epidermal stem cells (EpSCs) were cultured in a rotary cell culture system (RCCS) bioreactor to simulate microgravity. The metabolites of EpSCs were identified by liquid chromatography-mass spectrometry (LC-MS). Compared with normal gravity (NG) group, a total of 57 different metabolites of EpSCs were identified (P < 0.05, VIP > 1), including lipids and lipid-like molecules (51 molecules), amino acids (5 molecules), nucleosides, nucleotides, and analogues (1 molecule). According to the partial least squares discriminant analysis (PLS-DA) score plot, a VIP > 1 and P < 0.05 were obtained for the 57 different metabolites, of which 23 molecules were significantly downregulated and 34 were significantly upregulated in simulated microgravity (SMG) group. These results showed that SMG has a significant impact on different pathways, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis indicated that multiple pathways were involved, mainly the amino acid metabolism pathway, lipid metabolism pathway, membrane transport pathway, and cell growth and death pathways. Thus, the metabolic profile of EpSCs was changed under SMG. Exploring the metabolic profile of EpSCs would be helpful to further understand the growth characteristics of EpSCs under SMG, which will provide a new approach to explore the metabolomics mechanism of stress injury and repair trauma under SMG.
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9.
Intestinal Regeneration: Regulation by the Microenvironment.
Hageman, JH, Heinz, MC, Kretzschmar, K, van der Vaart, J, Clevers, H, Snippert, HJG
Developmental cell. 2020;(4):435-446
Abstract
Damage to the intestinal stem cell niche can result from mechanical stress, infections, chronic inflammation or cytotoxic therapies. Progenitor cells can compensate for insults to the stem cell population through dedifferentiation. The microenvironment modulates this regenerative response by influencing the activity of signaling pathways, including Wnt, Notch, and YAP/TAZ. For instance, mesenchymal cells and immune cells become more abundant after damage and secrete signaling molecules that promote the regenerative process. Furthermore, regeneration is influenced by the nutritional state, microbiome, and extracellular matrix. Here, we review how all these components cooperate to restore epithelial homeostasis in the intestine after injury.
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10.
Postnatal skeletal growth is driven by the epiphyseal stem cell niche: potential implications to pediatrics.
Chagin, AS, Newton, PT
Pediatric research. 2020;(6):986-990
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Abstract
Children's longitudinal growth is facilitated by the activity of the growth plates, cartilage discs located near the ends of the long-bones. In order to elongate these bones, growth plates must continuously generate chondrocytes. Two recent studies have demonstrated that there are stem cells and a stem cell niche in the growth plate, which govern the generation of chondrocytes during the postnatal growth period. The niche, which allows stem cells to renew, appears at the same time as the secondary ossification center (SOC) matures into a bone epiphysis. Thus, the mechanism of chondrocyte generation differs substantially between neonatal and postnatal age, i.e., before and after the formation of the mineralized epiphyses. Hence, at the neonatal age bone growth is based on a consumption of chondro-progenitors whereas postnatally it is based on the activity of the stem cell niche. Here we discuss potential implications of these observations in relation to longitudinal growth, including the effects of estrogens, nutrition and growth hormone.