Identification of Stromal Cells in Spleen Which Support Myelopoiesis

Abstract

Stromal cells in spleen organize tissue into red pulp, white pulp and marginal zone, and also interact with hematopoietic cells to regulate immune responses. This study has used phenotypic information of a previously described spleen stromal cell line called 5G3, which supports restricted hematopoiesis in vitro, to identify an equivalent stromal cell subset in vivo and to test its capacity to support hematopoiesis. Using stromal cell fractionation, phenotypic analysis, as well as cell growth and hematopoietic support assays, the Sca-1+gp38+Thy1.2+CD29+CD51+ fraction of spleen stroma has been identified as an equivalent stromal subset resembling the 5G3 cell counterpart. While heterogeneity may still exist within that subset, it has been shown to have superior hematopoietic support capacity compared with the 5G3 cell line, and all other spleen stromal cell fractions tested.

Targeting the Spleen as an Alternative Site for Hematopoiesis

“…It is now possible to perform stem cell isolation, single cell analysis, and to use in vitro differentiation protocols to identify the specific cell types that constitute stromal niches. Already we have embarked on analysis of splenic stromal subsets in mice and humans and have also developed protocols for engraftment of splenic stromal cells to form an ectopic or artificial niche (Figure ).…”

Niche mediated integrin signaling supports steady state hematopoiesis in spleen

“…Spleen stromal cell lines were later established that showed potenEal to support the development of hematopoieEc cells including immature dendriEc-like cells (39). Recently, these stromal cells that showed a rather restricted support of hematopoieEc processes were found equivalent to Sca-1 + gp38 + Thy1.2 + CD29 + CD51 + fracEon of spleen stroma (40). However, mechanisms that regulate the funcEon of spleen resident HSCs have not been uncovered thus far.…”

Time-Lapse Flow Cytometry: A Robust Tool to Assess Physiological Parameters Related to the Fertilizing Capability of Human Sperm

“…The E m fluorescence signal was calibrated by adding 1 µM valinomycin to induce PM hyperpolarization. Then, sequential additions of KCl were applied every 120 s to obtain final extracellular K + concentrations of 5, 10, 15, 25, and 40 mM [ 16 ], corresponding to theoretical E m values of −80, −63 −52 −40, and −28 mV, respectively. These theoretical E m values were obtained using the Nernst equation, assuming an intracellular K + concentration of 120 mM.…”

“…As a result, various efforts have been made to correlate the fertilizing capability of sperm with some of the physiological events that take place during sperm capacitation, a multistep phenomenon involving changes in sperm form and function [ 11 ], which is essential for fertilization, and which encompasses various molecular processes that sperm cells undergo during their transit inside the female reproductive tract [ 12 , 13 , 14 ]. Among these processes we can distinguish (a) changes in the plasma membrane’s ion permeability, which result in its hyperpolarization ([ 15 , 16 , 17 ], reviewed in [ 18 ]); (b) an increase in intracellular pH (pH i ) [ 19 , 20 , 21 ]; and (c) a rise in intracellular calcium concentration ([Ca 2+ ] i ) upon exposure to progesterone (Pg, a female hormone secreted by cumulus oophorus cells that surround the oocyte) [ 22 , 23 , 24 ]. These parameters have been independently analyzed in sperm samples from men with fertility issues, and results show that after capacitation induction, their sperm cells do not undergo pH i alkalinization, and they also fail to hyperpolarize (in some cases they depolarize) their plasma membrane (PM) [ 17 , 25 , 26 , 27 , 28 ].…”