Can TGF-β1 and rhBMP-2 act in synergy to transform bone marrow stem cells to discogenic-type cells?

Introduction  The recombinant human bone morphogenic protein-2 (rhBMP-2) is known to increase the proteoglycan production and chondrogenic gene expression in the disc cells. The transforming growth factor-beta 1 (TGF-β₁) can transform the bone marrow stem cells (BMDCs) into the disc-like cells. Materials and methods  We carried out an experiment to determine if TGF-β₁ and rhBMP-2 can act in synergy on BMDCs by increasing the production of sulfated-glycosaminoglycan (sGAG) and affecting the mRNA expression of aggrecan, type I collagen, and type II collagen. The BMDCs were isolated from the iliac crest and femur of a New Zealand white rabbit (1 year). The BMDCs were culured in monolayer and treated for 6 days with TGF-β₁ 10 ng/ml (group 1), rhBMP-2 200 ng/ml (group 2), and both TGF-β₁ 10 ng/ml and rhBMP-2 200 ng/ml (group 3: the combined group) in Dulbecco’s modified Eagle medium/F-12 with 1% fetal bovine serum. After 6 days, the sGAG content in the media was quantified using 1,9-dimethylmethylene blue staining and the mRNA expression of aggrecan, type I collagen, type II collagen, Sox-9, BMP-2, and BMP-7 were measured with the real-time PCR. The same BMDCs were also cultured in the chamber slide at 3 × 10⁴ cells/chamber. After 6 days treatment, the treated cells were immunofluorescence stained with aggrecan, type I collagen, type II collagen, anti-BMP-2, anti-BMP-7 antibodies. After that, we compared the number of positive immunofluorescence stained cells with fluorescence microscope. The sGAG production and mRNA expression for each group were normalized against the same parameters for a non-treatment group. Results and discussion  The sGAG production was increased 1.15*, 1.34*, and 1.45* times in the TGF-β₁ 10 ng/ml group, the rhBMP-2 200 ng/ml group, and the combined group respectively. The mRNA expression of aggrecan was increased 1.28, 3.42*, and 5.34* times, the mRNA expression of type I collagen was increased 0.86, 1.09, 1.17 times, the mRNA expression of type II collagen was increased 3.58*, 3.77*, and 10.78* times, the mRNA expression of Sox-9 was increased 1.29, 2.45, 2.75* times, the mRNA expression of BMP-2 was increased 1.14, 2.07, 4.43* times, and the mRNA expression of BMP-7 was increased 1.16, 1.49, 1.97* times, respectively for each group (* indicates p < 0.05). On the immunofluorescence staining of antibodies, the average positively immunofluorescence stained cells number for aggrecan were 4.2, 15.8*, 10*, and 22* according to the non-treatment group, TGF-β₁ 10 ng/ml group, rhBMP-2 200 ng/ml group, and the combined group respectively. The average positively immunofluorescence stained cells number for type I collagen were 7, 14.2*, 9.2*, 17.4* and the average positively immunofluorescence stained cells number for type II collagen were 8.5, 28.25*, 20.25*, 42.25* and the average positively immunofluorescence stained cells number for anti-BMP-2 were 5, 16.75*, 8.75*, 27.25* and the average positively immunofluorescence stained cells number for anti-BMP-7 were 3.25, 7.5*, 8.75*, 15.25* (* indicates p < 0.05). Conclusions  Both TGF-β₁ and rhBMP-2 alone, can increase proteoglycan production in the BMDCs. However, if they were used in combination, there is a synergistic effect. Similarly, the mRNA expressions of both aggrecan, type II collagen, Sox-9, BMP-2, and BMP-7 except for type I collagen were increased significantly when TGF-β₁ and rhBMP-2 were combined. The positive immunofluorescence stained cell numbers for aggrecan, type I, II collagen, BMP-2 and BMP-7 were also increased after each TGF-β₁ and rhBMP-2 treatment, and also more increased significantly in the aggrecan, type I, II collagen, BMP-2, and 7 when they were used jointly.     

Über angeborene Amputationen

Ohne Zusammenfassung Mit 3 Abbildungen im Text.     

Anti-tumor activity of heptaplatin in combination with 5-fluorouracil or paclitaxel against human head and neck cancer cells in vitro

Heptaplatin (HTP), a newly developed platinum analog, has been approved for the treatment of gastric cancers in South Korea. In this study we explored the potential of HTP for the treatment of head and neck squamous cell cancers (HNSCC). The anti-proliferative activity of HTP was evaluated in FaDu, a human HNSCC cell line. Combinations of HTP with 5-fluorouracil (5-FU) or paclitaxel (PTX) were determined using combination indexes, and were compared with combinations of cisplatin and 5-FU or PTX. In order to evaluate the transport of HTP into tumor tissue, its penetration through multicell layers (MCLs) of cancer cells was measured. Cisplatin+5-FU and HTP+5-FU showed additive to antagonistic interactions. In terms of the HTP+paclitaxel combination, HTP showed antagonism and additivity at the 50 and 80% growth inhibition levels, respectively. An additive interaction was obtained and apoptosis was increased by 2-fold at both inhibition levels when the combinatorial PTX dose was reduced to 1/10. HTP, but not cisplatin or oxaliplatin (L-OHP), maintained its anti-proliferative activity after MCL penetration at clinically relevant concentrations, which can be attributed to lower protein binding of HTP. In conclusion, the present study suggests that low-dose PTX may sensitize tumor cells to HTP. HTP also showed greater penetration through multilayers of tumor cells compared to cisplatin and L-OHP, which may be an important characteristic for solid tumor treatment. Overall, the present study supports the clinical development of HTP in combination with low-dose PTX against HNSCC.     

Life course social roles and women's health in mid‐life: causation or selection?

STUDY OBJECTIVE: To investigate whether relations between social roles and health are explained by health selection into employment and parenthood by examining the influence of early health on relations between long term social role histories and health in mid-life. DESIGN: Prospective, population based, birth cohort study. Participants and SETTING: Women from a national British cohort born in 1946, including 1171 women with a valid measure of self reported health at age 54 and valid work and family role measures at ages 26, 36, 43, and 53, as well as 1433 women with a valid body mass index (BMI) measure at age 53 and valid work and family role measures at ages 26, 36, 43, and 53. OUTCOME MEASURES: Self reported health at age 54 and obesity at age 53, taken from objective height and weight measures conducted by a survey nurse during face to face interviews in respondents' homes. MAIN RESULTS: Women who occupied multiple roles over the long term reported relatively good health at age 54 and this was not explained by early health. Women with weak long term ties to the labour market were more likely to be obese at age 53. Examination of body mass index (BMI) from age 15 showed that long term homemakers were larger than other women from age 26, but their mean BMI increased significantly more with age than that of other women. CONCLUSIONS: Relations between social roles and health were generally not explained by health selection into employment and parenthood, although some health selection may occur for obesity.     

Women's health in midlife: findings from a British birth cohort study

The Medical Research Council's National Survey of Health and Development (MRCNSHD) is a prospective cohort study of 2547 women and 2815 men, a sample of all the births that took place in England, Scotland and Wales between 3-9 March 1946. It is one of the longest running large-scale studies of human development in the world, aiming to identify lifetime biological, social and psychological pathways to health and disease, from early life to ageing. A special study of women's health in midlife and the menopausal transition in this cohort was undertaken by sending to women study members eight annual postal questionnaires from when they were 47 to when they were 54 years old. The findings from the women's health study so far have highlighted associations between multiple risk factors at each life stage, and women's health and disease in later life. The authors suggest that a life course approach may provide a better understanding of women's health during the middle years of life than an approach which restricts itself to contemporary social or hormonal experiences. Of particular interest are their results linking ovarian ageing to developmental factors. Replication of these findings in other life course cohorts is being sought.     

Birth weight,childhood size,and muscle strength in adult life: evidence from a birth cohort study

Environmental influences during gestation may have long-term effects on adult muscle strength. It is not known how early in adult life such effects are manifest and whether they are modified by childhood body size. The authors examined the relation between birth weight and hand grip strength in a prospective national birth cohort of 1,371 men and 1,404 women from the Medical Research Council National Survey of Health and Development who were aged 53 years in 1999. A positive relation between birth weight and adult grip strength remained after adjustment first for adult height and weight and then additionally for childhood height and weight (p = 0.006 for men and p = 0.01 for women). The effects of birth weight on grip strength did not vary by childhood or current body size and were not confounded by social class. To the authors' knowledge, this is the first study to show that birth weight has an important influence on muscle strength in midlife independent of later body size and social class. It suggests that birth weight is related to the number of muscle fibers established by birth and that even in middle age compensating hypertrophy may be inadequate. As the inevitable loss of muscle fibers proceeds in old age, a deficit in the number of fibers could threaten quality of life and independence.     

Activation-Induced Apoptosis of Peripheral Lymphocytes Treated with 7-Hydroxystaurosporine, UCN-01

7-hydroxystaurosporine (UCN-01) is a new anticancer agentwhich exerts an inhibitory effect on cell cycle check points andis currently under phase I clinical trials in US and Japan.Preliminary clinical data indicated that UCN-01 remained inplasma at high concentrations for long periods of time. Thisunavoidable high plasma drug exposure is likely to lead tohematological toxicities in patients. In the present study,cultured human peripheral blood lymphocytes (PBLs) were used toevaluate the possible hematological toxicities of UCN-01treatment. UCN-01 induces apoptosis, and the induction ofapoptosis-related surface markers were also examined toinvestigate the involvement of these molecules in UCN-01-inducedapoptosis in PBLs. in vitroviability of PBLs wasdecreased by high dose of UCN-01 (25 M, 3-day exposure). Thiseffect of UCN-01 was significantly suppressed by the presence ofhuman serum, suggesting that some specific inhibitory factor(s)in human serum may antagonize the lympholytic effect of UCN-01.The percentage of annexin V-positive PI-negative cells increasedwith exposure to UCN-01 in a time- and dose-dependent manner; byup to 30.3% after exposure to 25 M UCN-01 for 3 days.At the same time, the expression of both interleukin-2 receptor(IL-2R, CD25) and Fas (CD95), analyzed by flow cytometry, wasinduced. Con A-stimulated PBLs were more sensitive toUCN-01-induced apoptosis than non-stimulated lymphocytes andUCN-01 increased the sFas-L released into culture medium from conA-stimulated PBLs. Therefore, lymphocyte depletion mediated byactivation-induced apoptosis is likely to occur in patientstreated with UCN-01 at high doses.