In vivo 2D H NMR of mdx mouse muscle and myoblast cells during fusion: Evidence for a characteristic signal of long chain fatty acids

We have previously demonstrated that 2D ¹H NMR is suitable for studying cerebral metabolism. The same technique was used to study the hind leg muscle of normal (C57BL10) and dystrophic (mdx) mice. The results were compared to preliminary results for cultured muscle cells to determine the origin of fatty acid signals.     

EXPERIMENTAL SKELETAL MUSCLE GRAFTS AS A MODEL OF REGENERATION

Background : It is now well established that mature skeletal muscle has the ability to regenerate, and reports on this phenomenon have existed in the research literature for some 40 years. However, it is only relatively recently, largely due to the advances in microsurgery, that practising surgeons can make direct use of the regenerative ability of skeletal muscle. Methods : Most of the key data on skeletal muscle regeneration have come from experimental studies on muscle grafts in small animal models. One such model is the transplantation of the extensor digitorum muscle of the mouse or rat into the contralateral site, or the relocation of this muscle onto the surface of the tibialis anterior muscle. These and other models, together with the important cellular mechanisms involved in the regeneration of skeletal muscle, are reviewed briefly in this article. Results : Skeletal muscle cells regenerate rapidly in muscle grafts, arising from satellite cells in the surviving peripheral fibres of the graft within 2 days after grafting. The resultant myoblasts progress towards the necrotic graft centre and occupy the area by 5 days. Revascularization commences at 3 days after grafting, but reinnervation takes many weeks to complete. Conclusions : With the established knowledge on skeletal muscle regeneration, largely gained from experimental studies of muscle grafts, an understanding of these mechanisms should now be fundamental knowledge for today's practising surgeons.     

Cell–based cardiovascular repair

Cardiovascular cell therapy offers the first real potential to treat the underlying injuries associated with cardiac and vascular disease. By delivering appropriate exogenous cells to an injury site, the potential exists to mitigate injury or even to begin to reverse damage. Based on their inordinate pre–clinical promise as myogenic or angiogenic precursors, skeletal myoblasts and bone marrow or blood–derived mesenchymal and hematopoietic progenitor cells have all rapidly moved from bench to early clinical studies. From these parallel paths we are learning a number of useful lessons and have begun to visualize the hurdles to be overcome as we move these therapies forward. It is now obvious that cell–based cardiac and vascular repair are feasible—both early and later in the disease process. In fact, cell therapy may offer an unparalleled opportunity for improvement to millions of individuals living with cardiovascular disease. However, many questions about the technology remain. The mechanisms associated with cardiovascular repair remain unclear. Whether a best cell type, delivery method, or route of administration exists is unknown. And, whether cellbased disease prevention is feasible is still unanswerable. Now is the time to delve deeply into the questions of cell–based myocardial and vascular repair—even as we cautiously proceed clinically. Only by understanding these issues will we be able to decrease unanticipated clinical effects and to fulfill the potential promise of the most exciting opportunity yet to treat CVD. As we do so, we must prevent uncontrolled, poorly planned studies and until we understand cell therapy's potential, we must limit "too good to be true" promises. Only by addressing unanswered questions, carefully limiting our promises, and rigorously performing pre–clinical and clinical studies can we provide the surest opportunity for safely moving the field forward.     

周期性机械拉伸对C2C12成肌细胞增殖的影响

目的：探讨不同的周期性拉伸应变条件对C2C12成肌细胞增殖的影响。方法：周期性拉伸的各种力学条件通过BioFlex加载系统实现，采用应用流式细胞术和BrdU法对拉伸应变下的细胞增殖动力学变化进行分析，反映成肌细胞增殖情况。结果：不同的周期性机械拉伸条件影响C2C12细胞的增殖，拉伸的频率对C2C12细胞增殖有较大的影响。在0．5Hz拉伸频率下，2．5％、5％和10％的细胞变形幅度都不能促进细胞的增殖，其中10％（0．5Hz）的拉伸幅度抑制C2C12成肌细胞的增殖；而在0．125Hz拉伸频率下，10％的细胞变形幅度明显地促进C2C12成肌细胞的增殖。结论：较低的拉伸频率有利于成肌细胞的增殖，高频率的拉伸抑制成肌细胞的增殖。     

Improved angiogenic response in pig heart following ischaemic injury using human skeletal myoblast simultaneously expressing VEGF165 and angiopoietin-1

OBJECTIVE: To achieve angiogenic interaction between VEGF(165) and angiopoietin-1 (Ang-1) using a novel adenoviral bicistronic vector (Ad-Bic) encoding the two factors and delivered ex vivo using sex-mismatched human skeletal myoblasts. METHODS AND RESULTS: A myocardial infarction model was developed in 29 female pigs; randomised into four groups: DMEM (group-1, n=6); Adenovirus null (Ad-null) vector-myoblast (group-2, n=5); Ad-Ang-1 myoblast (group 3, n=7) and Ad-Bic-myoblast (group-4, n=11). Three weeks later, 5 ml DMEM without myoblasts or containing 3 x 10(8) myoblasts carrying lac-z gene and transduced with Ad-null, Ad-Ang-1 or Ad-Bic were injected intra-myocardially in and around the infarct. 2D-echocardiography and fluorescent microsphere studies 6- and 12-weeks post-treatment revealed significantly improved cardiac performance and regional blood flow in groups 3 and 4. Histological studies and Y-chromosome analysis revealed extensive survival of lac-z positive myoblasts staining positive for human proteins in the pig heart. ELISA, immunostaining and RT-PCR revealed that Ad-Bic transduced myoblasts concomitantly but transiently expressed hVEGF(165) and Ang-1 both in vitro and in vivo. Double fluorescent immunostaining of the tissue sections for vWFactor-III and smooth muscle actin showed significantly higher vascular density of mature blood vessels per low power microscopic field in groups 3 and 4 at 6- and 12-weeks. CONCLUSION: Our combined approach led to enhanced angiogenesis with a greater percentage of functionally mature blood vessels in a porcine heart.     

Primary myogenic cells see the light: improved survival of transplanted myogenic cells following low energy laser irradiation

BACKGROUND AND OBJECTIVES: There is a substantial need for finding new avenues to promote muscle recovery when acute skeletal muscle loss extends beyond the natural capacity of the muscle to recover. Maintenance and regeneration of skeletal muscles depend mainly on resident stem cells known as satellite cells. Nevertheless, there are situations in which a significant loss of muscle tissue exhausts the satellite cell pool. For such cases, cell therapy and tissue engineering are becoming promising alternatives. Thus far, attempts to supplement damaged host muscles with donor satellite cells by means of myoblast transplantation therapy were mostly unsuccessful due to massive and rapid loss of donor cells within few hours after transplantation. This study aims at following the effects of low-energy-laser irradiation on the fate of implanted myoblasts. STUDY DESIGN: Primary myogenic cells, harvested from male rat skeletal muscles, were irradiated with low energy laser, seeded on a biodegradable scaffold and expanded in vitro. The scaffold containing cells was transplanted into partially excised muscles of host female rats. Donor cells were identified in the host muscle tissue, using Y-chromosome in situ hybridization. RESULTS: In this study, we show that laser irradiated donor primary myogenic cells not only survive, but also fuse with host myoblasts to form a host-donor syncytium. CONCLUSIONS: Our data show that the use of low energy laser irradiation (LELI), a non-surgical tool, is a promising means to enhance both the survival and functionality of transplanted primary myogenic cells.     

软骨源性形态发生蛋白2基因转染自体成肌细胞表达软骨细胞表型的实验研究

目的研究软骨形态发生蛋白-2(CDMP-2)对小鼠成肌细胞向软骨分化的作用。方法体外培养小鼠成肌细胞,贴壁细胞传代,取第3代细胞,对照组(A组)以无血清L-DMEM培养液培养,实验组(B、C、D组)以无血清L-DMEM培养液培养,分别加入透明质酸钠、CDMP-2(100ng/mL)和CDMP-2基因转染的小鼠成肌细胞,14d后终止培养,倒置相差显微镜观察细胞形态,甲苯胺蓝染色糖胺聚糖(GAG),行Ⅱ型胶原免疫组织化学染色、灰度值分析和Western blot检测。结果培养14d后转染组和诱导组可见成肌细胞形态由梭形向软骨细胞的多边形转变。甲苯胺蓝染色示糖胺聚糖(GAGs)均匀分布于基质中,免疫组织化学染色和Western blot示实验组Ⅱ型胶原表达阳性,且CDMP-2基因转染组细胞表达Ⅱ型胶原水平高于单独给予CDMP-2,且具有统计学意义;对照组和透明质酸钠组未见阳性表达。结论 CDMP-2基因转染的小鼠成肌细胞能够更好的表达软骨特异性Ⅱ型胶原。