Objective To investigate the effect of macrophage inflammatory protein-1α (MIP-1α) on proliferation, migration and osteo-differentiation of human periodontal stem cells (hPDLSCs) and its possible mechanism. Methods A total of 16 healthy teeth (orthodontic minus premolar or blocked third molar) extracted from patients aged 12 to 25 years attending the outpatient clinic of the Department of Stomatology, the Second Affiliated Hospital of Xinjiang Medical University from November 2020 to October 2021 were collected and primary stem cells were cultured by tissue block method combined with enzymatic digestion, and the cell phenotype was identified by flow cytometry. (1) Cell biological characteristics experiment: the 3rd generation hPDLSCs were divided into 0 (control group), 1 and 10 μg/ml MIP-1α groups, and each group was then added with α-MEM medium containing volume fraction of 10% fetal bovine serum, 100 U/ml penicillin, and 2 mmol/L glutamine, respectively, and the proliferation ability of each group was detected by CCK-8 method after 24, 48 and 72 h of intervention. The lateral migration ability of cells in each group was detected by scratch assay after 24 h of intervention. (2) Effect and possible mechanism of osteo-differentiation: 3rd generation hPDLSCs were divided into 0 (control group) 1 and 10 μg/ml MIP-1α groups, and osteogenic induction solution was added to each group, and osteogenic ability of cells was detected by alkaline phosphatase (ALP) staining and semi-quantitative analysis 7 d after intervention and by alizarin red staining and semi-quantitative analysis 14 d after intervention;osteogenic ability of cells was detected by qRT-PCR and Western blotting. The mRNA and protein expression of osteogenic genes Runt-related transcription factor 2 (Runx2), bone bridge protein (OPN) and transcription factor SP7 (Osterix) and Notch1 receptor,Jagged 1 ligand and downstream factor Hey1 were detected by qRT-PCR and Western blotting 7 d after intervention. Results Flow cytometry results showed that hPDLSCs STRO-1 and CD146 showed positive expression, and CD34 showed negative expression.(1) In the experiment of cell biological characteristics, the results of CCK-8 method showed that the differences in OD values of hPDLSCs were not statistically significant (P>0.05) in 1 μg/ml and 10 μg/ml MIP-1α groups at 24 h and 48 h compared with control group; at 72 h, the differences in OD values of hPDLSCs were not statistically significant (P>0.05) in 1 μg/ml MIP-1α group compared with control group, while the differences in OD values of hPDLSCs in 10 μg/ml MIP-1α group were significantly higher (P<0.05). The results of scratch assay showed that the difference of scratch healing rate of cells in 1 μg/ml MIP-1α group was not statistically significant compared with that in control group (P>0.05), while the scratch healing rate of 10 μg/ml MIP-1α group was significantly higher (P<0.05). (2) In the experiments of the effect of osteo-differentiation and possible mechanism, ALP staining and semi-quantitative results showed that the ALP activity was obviously lower in 1 μg/ml and 10 μg/ml MIP-1α groups than that in control group (P<0.05). The results of alizarin red staining and semi-quantification showed that the number of mineralized nodules in 1 μg/ml and 10 μg/ml MIP-1α groups were significantly less than that in control group (P<0.05). qRT-PCR and Western blotting results showed that the osteogenesis-related genes Runx2, OPN and Osterix mRNA and protein expression levels in 1 μg/ml MIP-1α group of hPDLSCs were not statistically significant compared with control group, while those in the 10 μg/ml MIP-1α group were significantly lower (P<0.05). Notch1 mRNA and protein expression levels in 1 μg/ml and 10 μg/ml MIP-1α groups were significantly lower than those in control group (P<0.05); Jagged1 and Hey1 mRNA and protein expression levels in 10 μg/ml MIP-1α group were lower than those in control group (P<0.05). while the differences were not statistically significant in the 1 μg/ml MIP-1α group (P>0.05). Conclusion MIP-1α can promote proliferation and inhibit the osteo-differentiation of hPDLSCs, and the mechanism may be related to the inhibition of Notch signaling pathway activation.