Nicotine is one of the major chemical components of the cigarette smoke, which has been known as a risk factor for tendon ruptures including rotator cuff tears. This study investigated the effect of nicotine on tenocytes under cyclic‐stretched condition. Particularly, we focused on the morphologic changes of tenocytes and their expression of MMPs. Primary porcine tenocytes were obtained from the infraspinatus tendon. The cells were cultured on elastic chambers under static or cyclic‐stretched condition for 24 h in the existence of nicotine (0, 1, 10, and 100 µM). Cell shape, gene expression of collagen type I and III, MMPs (‐1, ‐2, ‐3, ‐9, and ‐13) and TIMPs (‐1, ‐2, and ‐3) and enzyme activity of MMP‐9 were analyzed using immunohistochemistry, RT‐PCR, and zymography. Tenocytes exposed to nicotine represented significantly decreased gene expressions in MMP‐9 (p < 0.001) and TIMP‐3 (p < 0.05) under the cyclic stretch. Enzymatic activity of MMP‐9 was also reduced by nicotine exposure in a dose‐dependent manner (p < 0.001). The down‐regulation of MMP and TIMP expression by nicotine shown in our in vitro experiment might deteriorate normal metabolism of the tendon. These mechanisms might affect the mechanical properties of the extracellular matrix of the rotator cuff tendon. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 645–650, 2013