scholarly journals Correction of the the liver morpho-functional state with stem cells in acute hepatitis

2020 ◽  
pp. 46-53
Author(s):  
И.Ю. Маклакова ◽  
Д.Ю. Гребнев
Keyword(s):  
Stem Cells ◽  
Carbon Tetrachloride ◽  
Hematopoietic Stem Cells ◽  
Toxic Hepatitis ◽  
Intracellular Regeneration ◽  
Hematopoietic Stem ◽  
Physiological Conditions ◽  
Intravenous Injections ◽  
And Control ◽  
Hepatic Protein Synthesis

Цель работы - изучение влияния сочетанной трансплантации мультипотентных мезенхимальных стромальных и гемопоэтических стволовых клеток на регенерацию печени в физиологических условиях и в условиях токсического гепатита. Методика. Эксперименты выполнены на 84 белых лабораторных мышах-самцах, возраст 7-8 мес. Токсический гепатит вызывали внутрибрюшинным введением четыреххлористого углерода (CCl4 в дозе 50 мкг/кг. Животным опытной группы в хвостовую вену вводили мультипотентные мезенхимальные стромальные (ММСК, 4 млн клеток/кг) и гемопоэтические стволовые клетки (ГСК, 330 тыс. клеток/кг). Для инъекции полученные клетки суспендировали в 0,2 мл 0,9 % раствора NaCl. Животным контрольной группы внутривенно вводили 0,2 мл 0,9 % раствора NaCl. Внутривенные введения осуществляли через 1 ч после введения четыреххлористого углерода однократно. Для трансплантации использовали ММСК третьего пассажа, а ГСК не подвергались культивированию. Изучали влияние сочетанной трансплантации ММСК и ГСК на биохимические показатели периферической крови и морфометрические показатели печени в физиологических условиях и после введения CCl4 на 1-е, 3-и, 7-е сут. Результаты. Показано, что проведение сочетанной трансплантации мультипотентных мезенхимальных стромальных и гемопоэтических стволовых клеток при токсическом гепатите приводило к снижению активности ферментов цитолиза, активации белоксинтезирующей функции печени. Также отмечалось увеличение количества гепатоцитов, повышение митотической активности, что указывает на активацию внутриклеточной регенерации. Обнаруженное увеличение числа двуядерных гепатоцитов, размеров ядра, ядерно-цитоплазматического соотношения говорит об усилении процесса внутриклеточной регенерации в печени. The aim of the work was studying the effect of combined transplantation of multipotent mesenchymal stromal and hematopoietic stem cells on regeneration of the liver under the physiological conditions and in toxic hepatitis. Methods. Experiments were performed on 84 white laboratory male mice aged 7-8 months. Toxic hepatitis was induced by administration of carbon tetrachloride (CCl4) at a dose of 50 µg/kg, i.p. Mice were divided into experimental and control groups. The experimental group received caudal vein injections of multipotent mesenchymal stromal (MMSC) and hematopoietic stem cells (HSC) derived from the placenta chorion of female mice at respective doses of 4M cells/kg and 330K cells/kg suspended in 0.2 ml of 0.9% NaCl. Control animals were given 0.2 ml of 0.9 % NaCl, i.v. Intravenous injections were performed once 1 hour following the administration of carbon tetrachloride. MMSCs of the third passage were used for transplantation, while transplanted HSCs were not cultured. Effects of the combined MMSC and HSC transplantation on blood biochemistry and liver morphometry were studied under the physiological conditions and at 1, 3, and 7 days after administration of CCl4. Results. In toxic hepatitis, the combined transplantation of multipotent mesenchymal stromal and hematopoietic stem cells resulted in decreased activity of cytolytic enzymes, activation of hepatic protein synthesis, increased number of hepatocytes, and increased mitotic activity indicative of activation of intracellular regeneration. Increases in the number of binuclear hepatocytes, nucleus size, and the nuclear-cytoplasmic ratio suggested an enhancement of intracellular regeneration in the liver.

Hematopoietic Stem Cells and Control of Hematopoiesis

2020 ◽  
pp. 67-108
Author(s):  
Mohamed Essawy ◽  
Ahmed Abdelfattah-Hassan ◽  
Eman Radwan ◽  
Mostafa F. Abdelhai ◽  
S. Elshaboury ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Hematopoietic Stem ◽  
And Control

Enhanced Engraftment of Human CD34+ Stem Cells in NOD/SCID Mice by Diprotin A Occurs by Inhibition of Recipient CD26/Dipeptidyl Peptidase-IV (DPP-IV).

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3178-3178
Author(s):  
Toshinao Kawai ◽  
Uimook Choi ◽  
Po-Ching Liu ◽  
Harry L. Malech
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Xenograft Model ◽  
Scid Mice ◽  
Control Group ◽  
Hematopoietic Stem ◽  
Human Cd34 ◽  
Dpp Iv ◽  
Cell Engraftment ◽  
And Control

Abstract CD26/DPP-IV (CD26) is a membrane-anchored ectoenzyme with N terminus exopeptidase activity that cleaves X-Pro-dipeptides. Stromal cell-derived factor 1 (SDF-1) and its receptor CXCR4 play a central role in trafficking of hematopoietic stem cells in the bone marrow. SDF-1 has a proline second from the N-terminus and is cleaved by CD26. Mouse hematopoietic progenitors express CD26 and a brief treatment of these cells with Diprotin A (Ile-Pro-Ile), a specific inhibitor of CD26, enhances engraftment. We examined the effect of Diprotin A treatment of peripheral blood human CD34+ stem cells (PBSC) with respect to subsequent responses to SDF-1 and with respect to engraftment in the NOD/SCID mouse xenograft model. We found that human CD34+ PBSC with colony forming potential are unlike mouse hematopoietic stem cells in that they lack the equivalent of CD26; and also are unlike mouse cells in that their response to SDF-1 and their engraftment in the NOD/SCID xenograft model are not affected by pre-treatment with Diprotin A. However, administration of Diprotin A intravenously to the NOD/SCID mouse at the time of transplant of human PBSC greatly enhances engraftment of the human PBSC, suggesting an effect primarily on the mouse stroma. Previous reports suggested that 70% of mouse lineage negative/sca-1 positive hematopoietic stem cells express CD26. However, freshly selected mobilized human CD34+ PBSC do not express detectable CD26, though after 4 days and 7 days of ex vivo culture in growth factors (SCF, flt3-ligand, TPO, IL3) 8.9% and 26.6% of cells express CD26, indicating that CD26 may only appear in later progenitors. At day 4 the cultured human PBSC were sorted by flow cytometry into CD26 positive and negative fractions. Only the CD26 negative fraction contained colony forming cells. 4 day-cultured human PBSC were exposed to Diprotin A 5mM for 15 minutes, washed and used in a filter transwell migration assay in response to SDF-1 at concentrations from 0.5 to 10 nM. There was no statistical difference between migration of Diprotin A treated and control PBSC, even in experiments with longer treatment with Diprotin A. When these human PBSC were transplanted into NOD/SCID mice there was no difference of engraftment between the Diprotin A treated group and control group. However, when 1x106 of 4 day-cultured PBSC were injected into NOD/SCID mice without or together with 2μmol of Diprotin A, there was a profound enhancement on subsequent engraftment in the group of mice injected with Diprotin A at time of transplant. At 6 weeks after transplantation the CD45+ human cell engraftment of the Diprotin A group was 6-fold increased compared to control group (49.6±8.2% vs. 8.1 ± 3.4%, p<.0001). Taken together with the colony assay, the in vitro migration studies, and lack of effect on engraftment when only the human PBSC are treated with Diprotin A, this result suggests that the enhanced engraftment of human PBSC in NOD/SCID mice is due to an action of Diprotin A on endogenous mouse CD26/DPP-IV (where the target is unknown, but possibly stromal cells). Although, further work is required to determine levels of expression of CD26/DPP-IV in human marrow stromal cells, it is possible to speculate that inhibitors of CD26/DPP-IV activity may provide a novel approach to improve stem cell engraftment in humans.

scholarly journals N-Cadherin in Osteolineage Cells Is Not Required for Maintenance of Hematopoietic Stem Cells

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2390-2390
Author(s):  
Adam Greenbaum ◽  
Daniel Link
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Conflicts Of Interest ◽  
Dominant Negative ◽  
Hematopoietic Stem ◽  
Conditional Deletion ◽  
Lineage Mapping ◽  
And Control ◽  
Osteoblast Lineage

Abstract Abstract 2390 Osteoblast lineage cells have been shown to play an important role in regulating hematopoietic stem cells (HSCs), and there is intense interest in identifying HSC regulatory molecules they produce. It has been reported that HSCs lie adjacent to spindle-shaped N-cadherin+ osteoblasts (SNO cells) and home to them in irradiated recipients, suggesting that N-cadherin may tether HSCs to their niche. Studies of N-cadherin expression in HSCs and its role in regulating HSC function have yielded conflicting results. Conditional deletion of Cdh2 (encoding N-cadherin) in HSCs had no affect on HSC number or function. On the other hand, silencing of N-cadherin using shRNA or expression of a dominant negative N-cadherin mutant resulted in the loss of HSC quiescence and repopulating activity. In addition to forming homodimers, N-cadherin is able to interact with other cadherins such E-cadherin, C-cadherin, and R-cadherin as well as non-cadherins such as KLRG1. Thus it is possible that expression of other cadherins in HSCs may compensate for the loss of N-cadherin. Rather than attempt to reconcile the conflicting results involving HSC production of N-cadherin, we chose to investigate what role that osteolineage production of N-cadherin plays in the regulation of hematopoiesis. Specifically, we conditionally deleted N-cadherin from osteoblast lineage cells using transgenic mice expressing Cre-recombinase under control of the osterix promoter (Osx-Cre mice). Our lineage mapping studies using the Osx-Cre mice demonstrated that this transgene directs recombination in SNO cells in the bone marrow. Accordingly, we intercrossed the Osx-Cre mice with Cdh2flox mice to generate N-cadherin-deleted (Cdh2flox/flox Osx-Cre) and control (Cdh2flox/flox) mice. N-cadherin expression was efficiently ablated in osteoblast lineage cells as assessed by mRNA expression (20-fold lower than control mice) and immunostaining of bone sections. Blood counts, bone marrow and spleen cellularity, and leukocyte differentials in N-cadherin-deleted mice were no different from control mice, indicating that basal hematopoiesis is normal. Moreover, the number of phenotypic HSCs (defined as lin− c-kit+ sca+ CD41− CD48− CD150+ cells) and their cell cycle status was normal. HSC long-term repopulating activity and self-renewal capacity were assessed by competitive repopulation assays and serial transplantation, respectively; we show that loss of osteoblast N-cadherin had no effect on these parameters. N-cadherin has been implicated in the homing and retention of HSCs to the bone marrow. However, we show that homing and engraftment of wildtype cells into N-cadherin-deleted recipients was normal. Finally, we tested the response to G-CSF, a potent HSC mobilizing stimulus, which leads to a profound loss of osteoblasts. N-cadherin-deleted mice showed normal mobilization of progenitors to the blood and spleen. Together, our data show that N-cadherin expression on SNO cells (and other osteoblast-lineage cells) is dispensable for HSC maintenance and trafficking. Disclosures: No relevant conflicts of interest to declare.

scholarly journals After migration into blood circulation, hematopoietic stem cells can stably self-renew and maintain bone marrow while being skewed toward myeloid lineage when submitted to serial transplantation

2021 ◽  
Author(s):  
Yasmine Even ◽  
Lin Yi ◽  
Chih-Kai Chang ◽  
Fabio M. Rossi
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Blood Circulation ◽  
Hematopoietic Stem ◽  
Hematological Disorders ◽  
Physiological Conditions ◽  
Therapeutic Research ◽  
Stimulating Factor ◽  
Serial Transplantation

AbstractHematopoietic stem cells (HSCs) mainly reside in bone marrow (BM) within niches providing an appropriate environment for their survival and self-renewal. Although, small numbers of HSCs can quit their residing environment to migrate into blood circulation and re-engraft elsewhere in BM. Mobilizing agents such as granulocyte colony stimulating factor (G-CSF) can amplify this process by inducing massive HSC mobilization into blood circulation. This method is widely used in clinics to treat hematological disorders. However, in physiological conditions, the properties of HSCs after migration (called migratory HSCs) remain incompletely characterized. In this study, we investigated the capacity of migratory HSCs to self-renew, reconstitute and maintain BM. We show that after migration, HSCs can stably self-renew and maintain BM in homeostasis. However, while stably repopulating BM of irradiated recipients, migratory HSCs show a defect in lymphoid lineage reconstitution when subjected to serial transplantations. Our findings provide interesting knowledge on HSC properties after migration, which may benefits therapeutic research on HSC-based therapies to treat hematological disorders.

scholarly journals Environmental Optimization Enables Maintenance of Quiescent Hematopoietic Stem Cells Ex Vivo

2018 ◽  
Author(s):  
Hiroshi Kobayashi ◽  
Takayuki Morikawa ◽  
Ayumi Okinaga ◽  
Fumie Hamano ◽  
Tomomi Hashidate-Yoshida ◽  
...  
Keyword(s):  
Stem Cells ◽  
Fatty Acid ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
High Fatty Acid ◽  
Quiescent State ◽  
Bone Marrow Niche ◽  
Hematopoietic Stem ◽  
Physiological Conditions

SUMMARYHematopoietic stem cells (HSCs) maintain lifelong hematopoiesis by remaining quiescent in the bone marrow niche. Recapitulation of a quiescent state in culture has not been achieved, as cells rapidly proliferate and differentiate in vitro. After exhaustive analysis of different environmental factor combinations and concentrations as a way to mimic physiological conditions, we were able to maintain engraftable quiescent HSCs for 1 month in culture under very low cytokine concentrations, hypoxia, and very high fatty acid levels. Exogenous fatty acids were required likely due to suppression of intrinsic fatty acid synthesis by hypoxia and low cytokine conditions. By contrast, high cytokine concentrations or normoxia induced HSC proliferation and differentiation. Our novel culture system provides a means to evaluate properties of steady state HSCs and test effects of defined factorsin vitrounder near-physiological conditions.

Hematopoietic Stem Cells in the Mouse Spleen

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2421-2421 ◽  
Author(s):  
Akiko Iseki ◽  
Yohei Morita ◽  
Hiromitsu Nakauchi ◽  
Hideo Ema
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Hematopoietic Stem Cells ◽  
Low Frequency ◽  
Extramedullary Hematopoiesis ◽  
Spleen Cells ◽  
Hematopoietic Stem ◽  
Hematopoietic Organ ◽  
Physiological Conditions ◽  
Cell Niche

Abstract The spleen is a hematopoietic organ in mice. Hematopoietic stem cells (HSCs) migrate into the spleen around embryonic day 14, and then migrate into the bone marrow (BM) around embryonic day 17. Thereafter, HSCs reside in both BM and spleen throughout the life of a mouse. The spleen is the major site of extramedullary hematopoiesis in pathological conditions. The spleen serves as an active hematopoietic organ in lethally irradiated mice for a while after transplantation with BM cells. Osteoblasts are considered to be one of the stem cell niche components. Because there are no osteoblasts in the spleen, niches in the spleen possibly functions differently from ones in the BM. The regulation of HSCs likely differs between the BM and spleen. In order to understand a role of spleen HSCs in physiological conditions, we have characterized HSCs in the spleen as compared with those in the BM. BM and spleen cells were obtained from 8–10 week-old C57BL/6 mice. Competitive repopulation showed that the repopulating activity per 106 BM cells was significantly greater than that per 106 spleen cells (about 10-fold). Limiting analysis showed that the frequency of long-term repopulating cells in BM cells was significantly higher than that in spleen cells (about 10-fold). As a result, the mean activity per BM stem cell was similar to that per spleen stem cell. Similarly to BM, CD34-negative, c-Kit-positive, Sca-1-positive, lineage markers-negative (CD34−KSL) cells were highly enriched in HSCs in the spleen. The frequency of CD34−KSL cells in the spleen was significantly lower than that in the BM. These data indicate that functionally equivalent HSCs exist in the spleen but at a low frequency. Data from single cell-transplantation supported this notion. The proportion of pyronin Y-negative G0 cells among BM CD34−KSL cells was greater than that among spleen CD34−KSL cells at any one time. BrdU-uptake analysis showed that spleen CD34− KSL cells were cycling more rapidly than BM CD34−KSL cells. These data suggest that spleen HSCs contribute to hematopoiesis to some extent under physiological conditions. BM and spleen HSCs may be interchangeable via the circulation. When BM HSCs are in the spleen, they are possibly under the control of spleen niches different from BM niches.

Cell staining for sorting of hematopoietic stem cells (HSC) and myeloid progenitors and isolating RNA from sorted cells

2006 ◽  
Author(s):  
Hideyo Hirai ◽  
Pu Zhang ◽  
Tajhal Dayaram ◽  
Christopher Hetherington ◽  
Shin-ichi Mizuno ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Hematopoietic Stem ◽  
Cell Staining ◽  
Myeloid Progenitors
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