The In Vitro Culture And Control Of Osteoblasts From Multiple Myeloma Patients

Objective:This study was to investigate the biological characteristics of osteoblasts cultured in vitro from bone marrow (BM) of multiple myeloma(MM) patients and to explore their generation and osteogenic potential. Then the affects by some factors such as bortezomib and MM patient serum on the osteogenic potential of osteoblasts from MM patients and normal subjects were observed.Methods:Fifteen MM patients and 10 healthy donors as controls were enrolled in this study. Osteoblasts from BM of MM patients were cultured in vitro. The morphology of these cells were observed, the growth pattern and double time were detected. The cultured cells were identified by ALP staining,Von Kossa staining and type I collagen immunohistochemistry assay. The generation and osteogenic potential of osteoblasts from MM patients and normal subjects were compared. The changes of their osteogenic potential and biological characteristics were observed. The antigens (CD34,CD138,CD45) on osteoblasts were examined with flow cytometric assay.The levels of IL-7 were measured with ELISA. The BMP2 mRNAs were measured by RT-PCR.Results:1. Osteoblasts from BM of MM patients could be cultured in vitro with dexamethasone,β-sodium glycerophosphate and vitamin C. The shape of the cells was almost spindle and polygon.Their double time was about 38 hours. ALP staining,Von Kossa staining and type I collagen immunohistochemistry staining were all positive to verify the osteoblasts. The quantity of osteoblasts from MM patients (5.77±1.91) was less than normal subjects (7.55±2.54) at the 6th day culture(p<0.05).There were calcium depositions in both groups after cultured for 4weeks culture. But the depositions of MM patients (6.12±1.63) was less than that of normal controls (15.83±2.23) (p＜O.05)2. The osteoblasts cultured with MM patient serum (7.38±1.13) were less than those without patient serum (8.25±2.59) (p<0.05). Bortezomib could not affect the cultured cells from normal person (p>0.05), but increased those from MM patients after 6days culter (8.94±2.09 vs 6.33±1.51) (p<0.05). After 4weeks culture, Von Kossa staining showed that there was more calcium depositions in MM ostesblasts with bortezomib (8.83±1.94) than those without bortezomib (6.12±1.63) (p<0.05), but less than those from normal controls. CD138, CD45, CD34 were not detected on the cultured cells. The level of IL-7 in serum of MM patients(2.07±0.71) was higher than that of normal controls(1.62±0.15)(p<0.05). The expression of BMP2 mRNA was seen in the normal osteoblasts and MM osteoblasts culured with bortezomib, but not seen in those without bortezomib.ConclusionsOsteoblasts could be cultured in vitro from BM of MM patients. The proliferation and osteogenic potential of osteoblasts from MM patients were decreased. Bortezomib was a positive regulatory of osteoblasts and MM patient serum was a negtive. They both could affect the proliferation and osteogenic potential of osteoblasts.