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Background: To investigate the effect of 3 months of Tai Chi Chuan (TCC) training on heart rate variability, blood lipids and cytokine production in elderly people. Methods: This was a longitudinal study with 3 months of follow-up. Participants were recruited from the community and divided randomly into two groups. The TCC trainees exercised the classical Yang's TCC for 40 minutes/session, seven times/week over a course of 3 months. Twenty-five TCC trainees (median age=57.0 years, range=50.0–67.0 years) and 25 control individuals (median age=53.0 years, range=46.5–58.5 years) were included in this study. Results: TCC training for 3 months significantly increased the fasting blood sugar (p<0.001), ratio of forced expiratory volume in the first second to the forced vital capacity (FEV1/FVC) (p<0.05), high-density lipoprotein-cholesterol (HDL-C) (p<0.05), tumor necrosis factor-α (TNF-α) (p<0.001), and interferon-γ (IFN-γ) (p<0.001) in the TCC trainees. In contrast, the total cholesterol (TC) and the ratio of TC to HDL-C (TC/HDL-C) of the TCC trainees were all significantly decreased after 3 months of TCC training (p<0.001). However, all heart rate variability (HRV) measures were not significantly changed after 3 months of TCC. Conclusion: Three months of TCC training can improve the pulmonary function, glucose availability and blood lipid profile, as well as increase the cytokine production, in middle-aged and elderly individuals. TCC may be beneficial to middle-aged and elderly people as a health-promoting calisthenics.