Despite numerous studies have demonstrated an association of type 2 diabetes mellitus (T2DM) and osteoporosis, the underlying mechanism connecting these two conditions remains elusive. Clinically, combined calcium and vitamin D supplement is the commonest osteoporosis therapy; however, recent studies have suggested an increase in cardiovascular risks associated with calcium plus vitamin D supplementation. Therefore, an alternative strategy in treating osteoporosis patients with T2DM is urgently needed. In this study, we hypothesized that combined administration of menaquinone-4 (vitamin K₂, biologically active form of vitamin K) and 1α,25-dihydroxyvitamin D₃ (vitamin D₃, biologically active form of vitamin D) as a novel therapy in treating osteoporosis of T2DM patients. Anabolic effect of vitamin K₂ and vitamin D₃, alone or in combination, was assessed on primary osteoblasts harvested from the iliac crests of C57BL/KsJ lean (db⁺/m⁺) and obese/diabetic (db⁺/db⁺, leptin receptor-deficient) mice. Furthermore, the underlying cellular mechanism was also investigated. Serum undercarboxylated osteocalcin (an indication of vitamin K₂ level) level was higher whereas vitamin D₃ level was lower in db⁺/db⁺ mice, and sections of the iliac crests of db⁺/db⁺ mice illustrated extensive porous structures filled with enlarged adipocytes compared with db⁺/m⁺ mice. Lower levels of bone anabolic markers and bone formation transcription factors (osteocalcin, Runx2, Dlx5, ATF4, type I collagen, OSX, alkaline phosphatase (ALP) activity, p-Smad1/5/8 and p-ERK1/2) were observed in the osteoblasts of db⁺/db⁺ mice. Acute vitamin D₃ (10 nM) application elicited a more sustained and greater magnitude of increase of [Ca²⁺]ᵢ in osteoblasts of db⁺/m⁺ mice when compared with db⁺/db⁺ mice. A significantly higher level of calcium deposits in osteoblasts was observed in db⁺/m⁺ mice when compared to db⁺/db⁺ mice. Co-administration of vitamin K₂ (10 nM) and vitamin D₃ (10 nM) caused an enhancement of calcium deposits in osteoblasts in both strains of mice. Vitamins K₂ and D₃ co-administration time-dependently (7, 14 and 21 days) increased the levels of bone anabolic markers and transcription factors for bone formation, with a greater magnitude of increase observed in osteoblasts of db⁺/db⁺ mice. Suppressed expression of calcium-sensing receptor (CaSR), F-actin, V-ATPase, vitamin D receptor (VDR) and pregnane X receptor (PXR) observed in osteoblasts of db⁺/db⁺ mice were partially reversed by combined vitamins treatment. Moreover, combined vitamins K₂ plus D₃ treatment significantly enhanced migration and the appearance of surface microvilli and ruffles of osteoblasts of db⁺/db⁺ mice. Effects of combined vitamins K₂ plus D₃ treatment observed in osteoblasts of db⁺/db⁺ and db⁺/m⁺ mice were eradicated by warfarin (20 µM, a vitamin K epoxide reductase inhibitor). Thus, our results illustrate that vitamins K₂ plus D₃ supplementation is a novel therapeutic strategy in treating osteoporosis of T2DM patients. / 儘管大量研究已證明第二類型糖尿病和骨質疏鬆症的關聯，連接這兩個病症的基本機制仍然是難以捉摸的。在臨床上，鈣和維生素D的綜合補充劑是最常見的骨質疏鬆症治療，然而最近的研究卻表明服用鈣和維生素D的綜合補充劑會增加患者的心血管風險，因此急切需要尋找可以給予同時患有骨質疏鬆症和第二類型糖尿病患者的替代治療。在本研究中，我們假設甲萘醌-4（維生素K₂，維生素K生物活性形式）和1α，25 - 二羥基維生素D₃（維生素D₃，維生素D的生物活性形式）可以嘗試在同時患有骨質疏鬆症和第二類型糖尿病患者身上作為一種革新的療法。本研究從C57BL/KsJ瘦削/非糖尿病 (db⁺/m⁺) 的小鼠和肥胖/帶有第二類型糖尿病基因 (db⁺/db⁺) 兼有瘦素受體缺陷的小鼠的髂嵴原始成骨細胞上對維生素K₂和維生素D₃單獨或組合使用的合成代謝作用進行了評估。此外，我們也對該成骨細胞的底層機制進行了一系列的研究。 / 在肥胖/帶有第二類型糖尿病基因的小鼠血清內低羧骨鈣素水平（維生素K₂水平的指標）較高而維生素D水平較低，另外，它們的髂嵴的部分與瘦削/非糖尿病的小鼠相比，呈現出比較廣泛的多孔結構並填滿了擴大的脂肪細胞。從肥胖/帶有第二類型糖尿病基因的小鼠的成骨細胞中，可以觀察到它們的骨合成代謝的標誌物和骨骼形成的轉錄因子 (骨鈣蛋白，Runx2，Dlx5，ATF4，第一類型骨膠原，OSX，鹼性磷酸酶 (ALP) 活性，p-Smad1/5/8和p-ERK1/2) 的水平比較低。急性維生素D₃ (10 nM) 的應用在瘦削/非糖尿病小鼠的成骨細胞比起在肥胖/帶有第二類型糖尿病基因的小鼠的成骨細胞引起更持續和更大幅度的細胞內鈣變化增加。在瘦削/非糖尿病小鼠的成骨細胞中比起在肥胖/帶有第二類型糖尿病基因的小鼠的成骨細胞有顯著較高的鈣沉積形成。維生素K₂ (10 nM) 和維生素D₃ (10 nM) 的綜合藥在兩種小鼠的成骨細胞中可以有效地增強鈣沉積的形成。維生素K₂和維生素D₃的綜合藥對增加骨合成代謝的標誌物和骨形成轉錄因子的水平有時間依賴性 (7，14和21日)，療程越長至21日，在肥胖/帶有第二類型糖尿病基因小鼠的成骨細胞中有更大的幅度的增加。合併維生素治療能部分有效地逆轉在肥胖/帶有第二類型糖尿病基因小鼠的成骨細胞中被抑制表達的鈣敏感受體 (CASR)，F-肌動蛋白，V-ATP酶，維生素D受體 (VDR) 和孕烷X受體 (PXR)。此外，結合維生素K₂加維生素D₃治療顯著增強了肥胖/帶有第二類型糖尿病基因小鼠的成骨細胞的細胞遷移和增加了成骨細胞表面外觀的微絨毛和褶皺。在瘦削/非糖尿病小鼠的成骨細胞及肥胖/帶有第二類型糖尿病基因的小鼠的成骨細胞上結合維生素K₂加維生素D₃的治療效果被華法林 (20 μM，維生素K環氧化物還原酶抑製劑) 根除。因此，我們的結果証明了維生素K₂加維生素D₃補充劑的結合使用可有效地作為治療第二類型糖尿病患者並患有骨質疏鬆症的一種新的治療策略。 / Poon, Chui Wa Christina. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2014.n5203 / Includes bibliographical references (leaves 135-151). / Abstracts also in Chinese. / Title from PDF title page (viewed on 26, October, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_1290651 |
| Date | January 2014 |
| Contributors | Poon, Chui Wa Christina (author.), Kwan, Yiu Wa (thesis advisor.), Chinese University of Hong Kong Graduate School. Division of Biomedical Sciences. (degree granting institution.) |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography, text |
| Format | electronic resource, electronic resource, remote, 1 online resource (xvi, 17-151 leaves) : illustrations, computer, online resource |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons "Attribution-NonCommercial-NoDerivatives 4.0 International" License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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