Objective To analyze the similarities and differences of bone microstructure and apoptosis phenotype of lateral column, middle column, and medial column in type L2 and L3 osteonecrosis of the femoral head (ONFH) specimens classified by China-Japan Friendship Hospital (CJFH) classification, so as to carry out a quantitative study of ONFH “three-columns structure theory” and to provide research support for the treatment of ONFH by rotational osteotomy through the base of femoral neck. MethodsDiscarded femoral head specimens from 16 patients (16 hips) with CJFH type L2 and L3 ONFH undergone total hip arthroplasty between April 2020 and February 2021 were selected as the research objects. First, the “three-column structure” of the femoral head was three-dimensionally segmented by Micro-CT, and the bone volume to total volume (BV/TV), bone surface area to bone volume ratio (BS/BV), trabecular spacing/separation (Tb.Sp), trabecular thickness (Tb.Th), and trabecular number (Tb.N) in the lateral column, middle column, and medial column were analyzed to compare the similarities and differences of parameters related to bone mass and trabecular structure among the three columns. Then, the specimens were cut with a oscillating saw and made into paraffin sections for HE staining and immunohistochemical staining of B-cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax). The differences of apoptosis phenotype between the three columns of ONFH samples of CJFH type L2 and L3 were evaluated by comparing the rate of empty lacunae and the rate of positive cells of immunohistochemical staining. ResultsThere were significant differences in BV/TV, Tb.Th, and Tb.N among the three columns of CJFH type L2 ONFH femoral head (P<0.05), with the largest in the medial column and the smallest in the lateral column; BS/BV and Tb.Sp of the lateral column were significantly greater than those of the medial column and middle column (P<0.05), no significant difference was found between the middle column and medial column (P>0.05). There was no significant difference in the bone parameters between the three columns of the CJFH type L3 ONFH femoral head (P>0.05). There was no significant difference in the rate of empty lacunae between the three columns of the CJFH type L2 and L3 ONFH femoral head (P>0.05). Immunohistochemical staining showed that a large number of tissue sections were detached, and only a small amount of non-specific staining was found in the sections without detachment, so the positive cell rate could not be calculated. Conclusion The middle and medial columns of the CJFH type L2 ONFH has better trabecular structures than the lateral column, and there is no significant difference in trabecular structures among the three columns of the CJFH type L3 ONFH.