Currently, the effect of drug treatment for osteoporosis is relatively poor, and the side effects are numerous and serious. Melatonin is a potential drug to improve bone mass in postmenopausal women. Unfortunately, the mechanism by which melatonin improves bone metabolism remains unclear. The aim of this study was to further investigate the potential mechanism of melatonin in the treatment of osteoporosis. The effects of melatonin on mitochondrial apoptosis protein, bmal1 gene, and related pathway proteins of RAW264.7 (mouse mononuclear macrophage leukaemia cells) were analyzed by western blot. Cell Counting Kit-8 was used to evaluate the effect of melatonin on cell viability. Flow cytometry was used to evaluate the effect of melatonin on the apoptosis of RAW264.7 cells and mitochondrial membrane potential. A reactive oxygen species (ROS) detection kit was used to evaluate the level of ROS in osteoclast precursors. We used bmal1-small interfering RNAs (siRNAs) to downregulate the Aims
Methods
Activated leukocyte cell adhesion molecule (ALCAM) has been shown to be involved in cell migration and in both homotypic/homophilic adhesion and heterotypic/heterophilic adhesion. It has been shown that a decreased level of ALCAM expression in human breast cancer tissue correlated with a significantly poor prognosis.
Primary breast cancer tissues (n=234) and non-neoplastic mammary tissue (n=34) were collected and patients were routinely followed up clinically after surgery. The immunohistochemical distribution and location of ALCAM was assessed in the normal breast tissue and carcinoma and the level of ALCAM transcripts in the frozen tissue was determined using real-time quantitative PCR. The results were analysed against the clinical data looking principally at the levels in patients with skeletal metastasis but also in relation to the nodal involvement, ER status, Nottingham Prognostic Index and survival. The immunohistochemical staining intensity shows that the cytoplasmic staining in normal breast tissue is significantly stronger than that in breast cancer tissue (p=0.023) and also the breast cancer tissue from patients who went onto develop skeletal metastasis (p=0.048). The ALCAM transcript levels were the lowest in patient with skeletal metastasis (p=0.0048) compared to those who were disease free. Significantly lower transcript levels were also found the patients who developed local recurrence (p=0.040), and who died from breast cancer (p= 0.0075). Other indicators of poor prognosis show a significant difference: patients with moderate and poor NPI prognosis lower levels than those with a good prognosis (p=0.05, p=0.0089 respectively); and lower in patients with a positive ER status than those ER negative patients (p=0.043). This study has for the first time shown that the patient who went on to develop skeletal metastasis tended to have the lowest levels of ALCAM transcript in their breast cancers. This fact could be used to provide patient with a more accurate prognosis and identify those who may benefit enhanced monitoring and early medical and orthopaedic treatment.
