探究一款以乳清蛋白(whey protein,WP)和亮氨酸(leucine,Leu)为主要功效成分的骨关节补充剂对关节炎大鼠软骨修复的可能机制。将40只SD大鼠分为空白组、关节炎模型组、阳性对照组、普通补充剂组、WP+Leu补充剂组,干预30 d后测定血清炎性因子、基质金属蛋白酶、软骨糖胺聚糖、软骨胶原蛋白,并对关节进行石蜡切片和番红O-固绿染色,在显微镜下观察。结果显示,关节炎大鼠血清促炎因子和基质金属蛋白酶水平显著上升,软骨糖胺聚糖和胶原蛋白含量显著降低,组织切片显示软骨出现明显损伤,符合早期关节炎症状。补充剂干预使血清促炎因子和基质金属蛋白酶水平显著降低,软骨糖胺聚糖和胶原蛋白含量显著回升,关节炎症状有所恢复,且WP和Leu联用效果要优于普通补充剂组。进一步研究发现,肌肉与软骨之间存在物理相互作用和内分泌相互串扰,WP和Leu干预后一方面通过增加肌肉量和肌肉力量,增强对软骨的力学刺激,上调软骨整合素β1水平,刺激磷脂酰肌醇三羟基激酶mRNA和蛋白激酶B mRNA转录,促进Ⅱ型胶原和蛋白聚糖的合成;另一方面提高血清成纤维细胞生长因子2(fibroblast growth factor-2, FGF-2)和胰岛素生长因子1水平,促进软骨细胞的合成代谢,同时上调FGF-2水平和性别决定区域迁移组框转录因子9 mRNA表达水平,促进间充质干细胞增值和向软骨细胞的分化。该研究显示在普通补充剂基础上添加WP和Leu可以通过增肌作用,借助肌肉和软骨相互作用帮助关节炎软骨修复,从而增强普通补充剂的功效,为相关产品开发提供理论依据。
This study aimed to investigate the effect of a bone and joint supplement mainly composed of whey protein (WP) and leucine (Leu) on the cartilage repair of rats with papain-induced osteoarthritis through musculoskeletal interaction.Forty SD rats were divided into a blank group, model group, positive control group, general supplement group, and WP+Leu supplement group.After 30 days of intervention, the serum inflammatory factors, matrix metalloproteinase, cartilage glycosaminoglycan, and cartilage collagen of rats were measured, and the joints were paraffin sections and safranin O-fast green staining was performed, and observed under a microscope.Results showed that the levels of serum proinflammatory factors and matrix metalloproteinases were significantly increased, the contents of cartilage glycosaminoglycan and collagen were significantly decreased, and the paraffin section showed obvious cartilage injury, which was consistent with early osteoarthritis symptoms.Supplement intervention significantly reduced the levels of proinflammatory factors and matrix metalloproteinases, significantly increased the contents of glycosaminoglycan and collagen in cartilage, and significantly recovered the symptoms of osteoarthritis, and WP and Leu could significantly improve the recovery effect.Further studies showed that physical interaction and endocrine cross-talk existed between muscle and cartilage.On the one hand, WP and Leu increasing muscle mass and muscle strength, and up-regulated the level of integrin-β1 by enhancing mechanical stimulation, then stimulated the transcription of phosphoinositide-3 kinase mRNA and protein kinase B mRNA, and promoted the synthesis of type Ⅱ collagen and proteoglycan.On the other hand, the serum fibroblast growth factor-2 and insulin growth factor-1 levels were increased to promote the anabolic metabolism of chondrocytes, and the level of fibroblast growth factor-2 and mRNA expression level of sex-determining region mobility group box 9 were up-regulated to promote the proliferation and differentiation of mesenchymal stem cells into chondrocytes.This study shows that the addition of WP and Leu on the basis of ordinary supplements can help arthritic cartilage repair through muscle building and muscle and cartilage interaction, thus enhancing the efficacy of ordinary supplements and providing theoretical basis for the development of related products.
[1] DE BRITO FONTANA H, HERZOG W.The role of muscles in knee joint osteoarthritis[J].Sports Orthopaedics and Traumatology, 2021, 37(2):85-100.
[2] SAWATSKY A, BOURNE D, HORISBERGER M, et al.Changes in patellofemoral joint contact pressures caused by vastus medialis muscle weakness[J].Clinical Biomechanics, 2012, 27(6):595-601.
[3] 金晶, 谢娇娇, 姜丹阳, 等.运动和细胞信号分子的交互作用及骨骼肌肥大调控机制[J].中国体育科技, 2021, 57(2):66-73.
JIN J, XIE J J, JIANG D Y, et al.Interaction between exercise and signaling molecules and regulation mechanism of skeletal muscle hypertrophy[J].China Sport Science and Technology, 2021, 57(2):66-73.
[4] BASSLEER C, ROVATI L, FRANCHIMONT P.Stimulation of proteoglycan production by glucosamine sulfate in chondrocytes isolated from human osteoarthritic articular cartilage in vitro[J].Osteoarthritis and Cartilage, 1998, 6(6):427-434.
[5] JOMPHE C, GABRIAC M, HALE T M, et al.Chondroitin sulfate inhibits the nuclear translocation of nuclear factor-kappaB in interleukin-1beta-stimulated chondrocytes[J].Basic & Clinical Pharmacology & Toxicology, 2008, 102(1):59-65.
[6] HURLEY M V.The role of muscle weakness in the pathogenesis of osteoarthritis[J].Rheumatic Disease Clinics of North America, 1999, 25(2):283-298.
[7] 张辉. 浅谈乳清蛋白与增肌运动[J].才智, 2009(33):297.
ZHANG H.Whey protein and muscle building exercise[J].Intelligence, 2009(33):297.
[8] 杨慧丽. 不同来源膳食蛋白质与慢性肾脏病发生及进展关系的研究进展[J].全科护理, 2022, 20(31):4359-4363.
YANG H L.Research progress on the relationship between different sources of dietary protein and the occurrence and progression of chronic kidney disease[J].Chinese General Practice Nursing, 2022, 20(31):4359-4363.
[9] DANGIN M, GUILLET C, GARCIA-RODENAS C, et al.The rate of protein digestion affects protein gain differently during aging in humans[J].The Journal of Physiology, 2003, 549(2):635-644.
[10] LEE J H, PAEK S H, SHIN H W, et al.Effect of fermented soybean products intake on the overall immune safety and function in mice[J].Journal of Veterinary Science, 2017, 18(1):25-32.
[11] D'ANTONA G, RAGNI M, CARDILE A, et al.Branched-chain amino acid supplementation promotes survival and supports cardiac and skeletal muscle mitochondrial biogenesis in middle-aged mice[J].Cell Metabolism, 2010, 12(4):362-372.
[12] KATSANOS C S, KOBAYASHI H, SHEFFIELD-MOORE M, et al.A high proportion of leucine is required for optimal stimulation of the rate of muscle protein synthesis by essential amino acids in the elderly[J].American Journal of Physiology.Endocrinology and Metabolism, 2006, 291(2):E381-E387.
[13] 伊薇特·查尔洛特·卢伊金, 乔治·韦尔兰, 马里昂·茹尔当.用于刺激肌肉蛋白合成的低热量高蛋白营养组合物:中国,CN102665736B[P].2014-10-29.
CHARLOT-LUYKIN Y, VERLAND G, JOURDAN M.A low-calorie, high-protein nutrient composition for stimulating muscle protein synthesis:China, CN102665736B[P].2014-10-29.
[14] MURRAY D G.Experimentally induced arthritis using intra-articular papain[J].Arthritis & Rheumatism, 1964, 7(3):211-219.
[15] MANKIN H J, LIPPIELLO L.Biochemical and metabolic abnormalities in articular cartilage from osteo-arthritic human hips[J].The Journal of Bone and Joint Surgery.American Volume, 1970, 52(3):424-434.
[16] OH M J, CHOI H D, HA S K, et al.Immunomodulatory effects of polysaccharide fraction isolated from Fagopyrum esculentum on innate immune system[J].Biochemical and Biophysical Research Communications, 2018, 496(4):1210-1216.
[17] 刘映岐. 菟丝子治疗膝骨性关节炎实验及临床验证研究[D].成都:成都中医药大学, 2017.
LIU Y Q.Experimental and clinical validation of dodder in the treatment of knee osteoarthritis[D].Chengdu:Chengdu University of TCM, 2017.
[18] HALLER J M, SWEARINGEN C A, PARTRIDGE D, et al.Intraarticular matrix metalloproteinases and aggrecan degradation are elevated after articular fracture[J].Clinical Orthopaedics and Related Research, 2015, 473(10):3280-3288.
[19] 文军, 荣晓凤.复方大黄散对类风湿关节炎大鼠MMP-10及IL-33的影响[J].免疫学杂志, 2015, 31(1):41-46.
WEN J, RONG X F.Effects of the combination of compound rhubarb powder and methotrexate on the expression of MMP-10 and IL-33 in rheumatoid arthritis rats[J].Immunological Journal, 2015, 31(1):41-46.
[20] BOKHARI R A, TANTOWI N A C A, LAU S F, et al.Java tea (Orthosiphon stamineus) protected against osteoarthritis by mitigating inflammation and cartilage degradation:A preclinical study[J].Inflammopharmacology, 2018, 26(4):939-949.
[21] 王也, 张力木, 齐帅, 等.IL-6、IL-33、IL-10在牙周炎和类风湿关节炎中的表达和相关性分析[J].上海口腔医学, 2021, 30(5):498-503.
WANG Y, ZHANG L M, QI S, et al.Preliminary study on the correlation between IL-6, IL-33, and IL-10 and periodontitis and rheumatoid arthritis[J].Shanghai Journal of Stomatology, 2021, 30(5):498-503.
[22] SCHWARZ U S, ERDMANN T, BISCHOFS I B.Focal adhesions as mechanosensors:The two-spring model[J].Biosystems, 2006, 83(2-3):225-232.
[23] CHONG K W, CHANALARIS A, BURLEIGH A, et al.Fibroblast growth factor 2 drives changes in gene expression following injury to murine cartilage in vitro and in vivo[J].Arthritis & Rheumatism, 2013, 65(9):2346-2355.
[24] COUTTS R D, SAH R L, AMIEL D.Effects of growth factors on cartilage repair[J].Instructional Course Lectures, 1997, 46:487-494.
[25] HANDORF A M, LI W J.Fibroblast growth factor-2 primes human mesenchymal stem cells for enhanced chondrogenesis[J].PLoS One, 2011, 6(7):e22887.
[26] KIM S H, TURNBULL J, GUIMOND S.Extracellular matrix and cell signalling:The dynamic cooperation of integrin, proteoglycan and growth factor receptor[J].Journal of Endocrinology, 2011, 209(2):139-151.
[27] 梁功强, 李富林, 程锦文, 等.膝关节炎患者关节腔药物注射方案标准化现状及评价[J].中国标准化, 2023(6):224-228.
LIANG G Q, LI F L, CHENG J W, et al.Current situation and evaluation of standardization of intra-articular drug injection scheme for patients with knee osteoarthritis[J].China Standardization, 2023(6):224-228.
[28] VARGHESE S, THEPRUNGSIRIKUL P, SAHANI S, et al.Glucosamine modulates chondrocyte proliferation, matrix synthesis, and gene expression[J].Osteoarthritis and Cartilage, 2007, 15(1):59-68.
[29] UITTERLINDEN E J, KOEVOET J L M, VERKOELEN C F, et al.Glucosamine increases hyaluronic acid production in human osteoarthritic synovium explants[J].BMC Musculoskeletal Disorders, 2008, 9:120.
