Direct differentiation of rat skin fibroblasts into cardiomyocytes.

Myocardial infarction (MI) is one of the major cardiovascular diseases caused by diminished supply of nutrients and oxygen to the heart due to obstruction of the coronary artery. Different treatment options are available for cardiac diseases, however, they do not completely repair the damage. Therefore, reprogramming terminally differentiated fibroblasts using transcription factors is a promising strategy to differentiate them into cardiac like cells in vitro and to increase functional cardiomyocytes and reduce fibrotic scar in vivo. In this study, skin fibroblasts were selected for reprogramming because they serve as a convenient source for the autologous cell therapy. Fibroblasts were isolated from skin of rat pups, propagated, and directly reprogrammed towards cardiac lineage. For reprogramming, two different approaches were adopted, i.e., cells were transfected with: (1) combination of cardiac transcription factors; GATA4, MEF2c, Nkx2.5 (GMN), and (2) combination of cardiac transcription factors; GATA4, MEF2c, Nkx2.5, and iPSC factors; Oct4, Klf4, Sox2 and cMyc (GMNO). After 72 h of transfection, cells were analyzed for the expression of cardiac markers at the mRNA and protein levels. For in vivo study, rat MI models were developed by ligating the left anterior descending coronary artery and the reprogrammed cells were transplanted in the infarcted heart. qPCR results showed that the reprogrammed cells exhibited significant upregulation of cardiac genes. Immunocytochemistry analysis further confirmed cardiomyogenic differentiation of the reprogrammed cells. For the assessment of cardiac function, animals were analyzed via echocardiography after 2 and 4 weeks of cell transplantation. Echocardiographic results showed that the hearts transplanted with the reprogrammed cells improved ejection fraction, fractional shortening, left ventricular internal systolic and diastolic dimensions, and end systolic and diastolic volumes. After 4 weeks of cell transplantation, heart tissues were harvested and processed for histology. The histological analysis showed that the reprogrammed cells improved wall thickness of left ventricle and reduced fibrosis significantly as compared to the control. It is concluded from the study that novel combination of cardiac transcription factors directly reprogrammed skin fibroblasts and differentiated them into cardiomyocytes. These differentiated cells showed cardiomyogenic characters in vitro, and reduced fibrosis and improved cardiac function in vivo. Furthermore, direct reprogramming of fibroblasts transfected with cardiac transcription factors showed better regeneration of the injured myocardium and improved cardiac function as compared to the indirect approach in which combination of cardiac and iPSC factors were used. The study after further optimization could be used as a better strategy for cell-based therapeutic approaches for cardiovascular diseases.

Subsurface skin renewal by treatment with a 1450-nm laser in combination with dynamic cooling.

A new nonablative laser device, Smoothbeam, has been under evaluation for nonablative wrinkle reduction in skin with minimal side effects. This device incorporates a laser at 1450-nm wavelength to heat the dermis and cryogen spray cooling to prevent epidermal damage. The thermal injury created is internal and imperceptible. The wound-healing response to this internal injury causes improvement in the appearance of skin wrinkles. Biopsies taken immediately after treatment showed mild residual thermal damage (RTD) at a depth range of 150 to 400 microm, which is the dermal zone where most solar elastosis resides. Biopsies from two months after treatment showed fibroplasia extending over a range of depths similar to the acute RTD zones. An improvement in wrinkle severity was noted on the treated side compared with the control side.

Acne treatment with a 1,450 nm wavelength laser and cryogen spray cooling.

BACKGROUND AND OBJECTIVES: A laser with a wavelength in the mid-IR range targeting the depth in skin where sebaceous glands are located in combination with cryogen spray cooling was evaluated for treatment of acne. In this non-ablative treatment, the laser energy heats the dermal volume encompassing sebaceous glands whereas the cold cryogen spray preserves the epidermis from thermal damage. STUDY DESIGN/MATERIALS AND METHODS: Monte Carlo simulations and heat transfer calculations were performed to optimize the heating and cooling parameters. A variety of heating and cooling parameters were tested in an in vivo rabbit ear study to evaluate the histological effect of the device on sebaceous glands and skin. Similar experiments were performed on ex vivo human skin. A clinical study for the treatment of acne on backs of human males was also conducted. RESULTS: Monte Carlo simulations and heat transfer calculations resulted in a thermal damage profile that showed epidermal preservation and peak damage in the upper dermis where sebaceous glands are located. Ex vivo human skin histology confirmed the damage profile qualitatively. In vivo rabbit ear histology studies indicated short-term thermal alteration of sebaceous glands with epidermal preservation. In the human clinical study on the back, a statistically significant reduction in lesion count on the treated side compared to the control side was seen (p < 0.001). Side effects were transient and few. CONCLUSIONS: The studies reported here demonstrate the feasibility of treating acne using a photothermal approach with a mid-IR laser and cryogen cooling.