Person:

Zhou, Shuanhu

1,25-Dihydroxyvitamin D3 [1,(25(OH){2}D{3})] has many noncalcemic actions that rest on inhibition of proliferation and promotion of differentiation in malignant and normal cell types. 1,(25(OH){2}D{3}) stimulates osteoblast differentiation of human marrow stromal cells (hMSCs), but little is known about the effects of 25-hydroxyvitamin D3 [(25(OH)D_{3})] on these cells. Recent evidence shows that hMSCs participate in vitamin D metabolism and can activate (25(OH)D_{3}) by CYP27B1/1(\alpha)-hydroxylase. These studies test the hypothesis that antiproliferative and prodifferentiation effects of (25(OH)D_{3}) in hMSCs depend on CYP27B1. We studied hMSCs that constitutively express high (hMSCs(^{hi-1\ \alpha})) or low (hMSCs(^{lo-1\ \alpha})) levels of CYP27B1 with equivalent expression of CYP24A1 and vitamin D receptor. In hMSCs(^{hi-1\ \alpha}), (25(OH)D_{3}) reduced proliferation, downregulated proliferating cell nuclear antigen (PCNA), upregulated p21(^{Waf1/Cip1}), and decreased cyclin D1. Unlike 1,(25(OH){2}D{3}), the antiapoptotic effects of (25(OH)D_{3}) on Bax and Bcl-2 were blocked by the P450 inhibitor ketoconazole. The antiproliferative effects of (25(OH)D_{3}) in hMSCs(^{hi-1\ \alpha}) and of 1,(25(OH){2}D{3}) in both samples of hMSCs were explained by cell cycle arrest, not by increased apoptosis. Stimulation of osteoblast differentiation in hMSCs(^{hi-1\ \alpha}) by (25(OH)D_{3}) was prevented by ketoconazole and upon transfection with CYP27B1 siRNA. These data indicate that CYP27B1 is required for (25(OH)D_3)'s action in hMSCs. Three lines of evidence indicate that CYP27B1 is required for the antiproliferative and prodifferentiation effects of (25(OH)D_{3}) on hMSCs: Those effects were not seen (1) in hMSCs with low constitutive expression of CYP27B1, (2) in hMSCs treated with ketoconazole, and (3) in hMSCs in which CYP27B1 expression was silenced. Osteoblast differentiation and skeletal homeostasis may be regulated by autocrine/paracrine actions of (25(OH)D_{3}) in hMSCs.

Stem cell function decline during ageing can involve both cell intrinsic and extrinsic mechanisms. Bone and blood formation are intertwined in bone marrow, therefore haematopoietic cells and bone cells could be extrinsic factors for each other. In this study, we assessed the paracrine effects of extrinsic factors from haematopoietic cells on human mesenchymal stem cells (MSCs). Our data showed that haematopoietic cells stimulate proliferation, osteoblast differentiation and inhibit senescence of MSCs; TNF-α, PDGF-β, Wnt1, 4, 6, 7a and 10a, sFRP-3 and sFRP-5 are dominantly expressed in haematopoietic cells; the age-related increase of TNF-α in haematopoietic cells may perform as a negative factor in the interactions of haematopoietic cells on MSCs via TNF-α receptors and then activating NF-κB signaling or Wnt/β-catenin signaling to induce senescence and reduce osteoblast differentiation in MSCs. In conclusion, our data demonstrated that there are paracrine interactions of haematopoietic cells on human MSCs; immunosenescence may be one of the extrinsic mechanisms by which skeletal stem cell function decline during human skeletal ageing.

Subarachnoid hemorrhage (SAH) results in significant nerve dysfunction, such as hemiplegia, mood disorders, cognitive and memory impairment. Currently, no clear measures can reduce brain nerve damage. The study of brain nerve protection after SAH is of great significance. We aim to evaluate the protective effects and the possible mechanism of methazolamide in C57BL/6J SAH animal model in vivo and in blood-induced primary cortical neuron (PCNs) cellular model of SAH in vitro. We demonstrate that methazolamide accelerates the recovery of neurological damage, effectively relieves cerebral edema, and improves cognitive function in SAH mice as well as offers neuroprotection in blood- or hemoglobin-treated PCNs and partially restores normal neuronal morphology. In addition, western blot analyses show obviously decreased expression of active caspase-3 in methazolamide-treated SAH mice comparing with vehicle-treated SAH animals. Furthermore, methazolamide effectively inhibits ROS production in PCNs induced by blood exposure or hemoglobin insult. However, methazolamide has no protective effects in morality, fluctuation of cerebral blood flow, SAH grade, and cerebral vasospasm of SAH mice. Given methazolamide, a potent carbonic anhydrase inhibitor, can penetrate the blood–brain barrier and has been used in clinic in the treatment of ocular conditions, it provides potential as a novel therapy for SAH.