Diamond encapsulated photovoltaics for transdermal power delivery.

A safe, compact and robust means of wireless energy transfer across the skin barrier is a key requirement for implantable electronic devices. One possible approach is photovoltaic (PV) energy delivery using optical illumination at near infrared (NIR) wavelengths, to which the skin is highly transparent. In the work presented here, a subcutaneously implantable silicon PV cell, operated in conjunction with an external NIR laser diode, is developed as a power delivery system. The biocompatibility and long-term biostability of the implantable PV is ensured through the use of an hermetic container, comprising a transparent diamond capsule and platinum wire feedthroughs. A wavelength of 980 nm is identified as the optimum operating point based on the PV cell's external quantum efficiency, the skin's transmission spectrum, and the wavelength dependent safe exposure limit of the skin. In bench-top experiments using an external illumination intensity of 0.7 W/cm(2), a peak output power of 2.7 mW is delivered to the implant with an active PV cell dimension of 1.5 × 1.5 × 0.06 mm(3). This corresponds to a volumetric power output density of ~20 mW/mm(3), significantly higher than power densities achievable using inductively coupled coil-based approaches used in other medical implant systems. This approach paves the way for further ministration of bionic implants.

Survival of rat functional dental pulp cells in vascularized tissue engineering chambers.

Regenerative endodontics aims to preserve, repair or regenerate the dental pulp tissue. Dental pulp stem cells, have a potential use in dental tissue generation. However, specific requirements to drive the dental tissue generation are still obscured. We established an in vivo model for studying the survival of dental pulp cells (DPC) and their potential to generate dental pulp tissue. DPC were mixed with collagen scaffold with or without slow release bone morphogenic protein 4 (BMP-4) and fibroblast growth factor 2 (FGF2). The cell suspension was transplanted into a vascularized tissue engineering chamber in the rat groin. Tissue constructs were harvested after 2, 4, 6, and 8 weeks and processed for histomorphological and immunohistochemical analysis. After 2 weeks newly formed tissue with new blood vessel formation were observed inside the chamber. DPC were found around dentin, particularly around the vascular pedicle and also close to the gelatin microspheres. Cell survival, was confirmed up to 8 weeks after transplantation. Dentin Sialophosphoprotein (DSPP) positive matrix production was detected in the chamber, indicating functionality of dental pulp progenitor cells. This study demonstrates the potential of our tissue engineering model to study rat dental pulp cells and their behavior in dental pulp regeneration, for future development of an alternative treatment using these techniques.

Myogel, a novel, basement membrane-rich, extracellular matrix derived from skeletal muscle, is highly adipogenic in vivo and in vitro.

BACKGROUND/AIMS: Biological and synthetic scaffolds play important roles in tissue engineering and are being developed towards human clinical applications. Based on previous work from our laboratory, we propose that extracellular matrices from skeletal muscle could be developed for adipose tissue engineering. METHODS: Extracellular matrices (Myogels) extracted from skeletal muscle of various species were assessed using biochemical assays including ELISA and Western blotting. Biofunctionality was assessed using an in vitro differentiation assay and a tissue engineering construct model in the rat. RESULTS: Myogels were successfully extracted from mice, rats, pigs and humans. Myogels contained significant levels of laminin alpha4- and alpha2-subunits and collagen I compared to Matrigel, which contains laminin 1 (alpha1beta1gamma1) and collagen IV. Levels of growth factors such as fibroblast growth factor 2 were significantly higher than Matrigel, vascular endothelial growth factor-A levels were significantly lower and all other growth factors were comparable. Myogels reproducibly stimulated adipogenic differentiation of preadipocytes in vitro and the growth of adipose tissue in the rat. CONCLUSIONS: We found Myogel induces adipocyte differentiation in vitroand shows strong adipogenic potential in vivo, inducing the growth of well-vascularised adipose tissue. Myogel offers an alternative for current support scaffolds in adipose tissue engineering, allowing the scaling up of animal models towards clinical adipose tissue engineering applications.

Zymosan-induced inflammation stimulates neo-adipogenesis.

OBJECTIVE: To investigate the potential of inflammation to induce new adipose tissue formation in the in vivo environment. METHODS AND RESULTS: Using an established model of in vivo adipogenesis, a silicone chamber containing a Matrigel and fibroblast growth factor 2 (1 microg/ml) matrix was implanted into each groin of an adult male C57Bl6 mouse and vascularized with the inferior epigastric vessels. Sterile inflammation was induced in one of the two chambers by suspending Zymosan-A (ZA) (200-0.02 microg/ml) in the matrix at implantation. Adipose tissue formation was assessed at 6, 8, 12 and 24 weeks. ZA induced significant adipogenesis in an inverse dose-dependent manner (P<0.001). At 6 weeks adipose tissue formation was greatest with the lowest concentrations of ZA and least with the highest. Adipogenesis occurred both locally in the chamber containing ZA and in the ZA-free chamber in the contralateral groin of the same animal. ZA induced a systemic inflammatory response characterized by elevated serum tumour necrosis factor-alpha levels at early time points. Aminoguanidine (40 microg/ml) inhibited the adipogenic response to ZA-induced inflammation. Adipose tissue formed in response to ZA remained stable for 24 weeks, even when exposed to the normal tissue environment. CONCLUSIONS: These results demonstrate that inflammation can drive neo-adipogenesis in vivo. This suggests the existence of a positive feedback mechanism in obesity, whereby the state of chronic, low-grade inflammation, characteristic of the condition, may promote further adipogenesis. The mobilization and recruitment of a circulating population of adipose precursor cells is likely to be implicated in this mechanism.

Smooth muscle alpha actin and myosin heavy chain expression in the vascular smooth muscle cells surrounding human endometrial arterioles.

Endometrial spiral arterioles are believed to play a major role in controlling menstruation. These arterioles coil and grow through the secretory stages of the cycle, unlike the 'straight' endometrial arterioles that remain uncoiled. We postulate that alterations in the growth and development of spiral arterioles, in particular the vascular smooth muscle cells (VSMC), may contribute to menorrhagia. We examined smooth muscle alpha actin (alphaSMA) and myosin heavy chains (MHC), two VSMC differentiation markers, in the endometrial arterioles of 64 women, comparing them in controls, menorrhagic tissues and across the menstrual cycle. alphaSMA and MHC expression were determined immunohistochemically then evaluated using computer-aided image analysis. alphaSMA expression in the straight arterioles of menorrhagic women was reduced in the early secretory stage of the cycle and significantly decreased at the mid-secretory stage of the cycle (0.67 +/- 0.03 versus 0.55 +/- 0.04, P

Vascular smooth muscle cell proliferation in arterioles of the human endometrium.

Menorrhagia affects approximately 15% of all women, often without identifiable cause. Endometrial spiral arterioles are believed to play a major role in controlling menstruation, and are a major site of menstrual loss. We postulate that alterations in the growth and development of spiral arterioles, particularly the vascular smooth muscle cells (VSMC), may contribute to menorrhagia. We examined VSMC proliferation around endometrial arterioles in control and menorrhagic tissues and the possible roles of transforming growth factor beta (TGF-beta) and endothelin in this process. Proliferating VSMC were located immuno-histochemically, then evaluated using computer-aided image analysis. VSMC proliferation was low and constant during the early stages of the menstrual cycle, increasing at the mid to late secretory stages (P < 0.002). Menorrhagic women had significantly reduced VSMC proliferation in their spiral arterioles at the mid and late secretory stages (P < 0.02). VSMC around straight arterioles proliferated at similar rates across the cycle, apart from a significant decrease in VSMC proliferation in menorrhagic women at the late secretory stage (P < 0.002). Endothelin concentrations decreased significantly in the epithelium of menorrhagic women (P = 0.05), while TGF-beta demonstrated no significant differences in the mid to late secretory tissues studied. The results indicate a significant functional difference between the spiral arterioles of control and menorrhagic women that may play a role in menorrhagia, while leaving the roles of endothelin and TGF-beta undetermined.

Vascular smooth muscle alpha-actin distribution around endometrial arterioles during the menstrual cycle: increased expression during the perimenopause and lack of correlation with menorrhagia.

Menorrhagia affects approximately 9% of all women, increasing to 20% during the perimenopause. The majority of menstrual loss occurs through the spiral arterioles - specialized endometrial vessels that are intimately involved in controlling menstruation. Our aim was to compare the distribution of vascular smooth muscle alpha-actin using immunohistochemical techniques in the endometrium of women before and during the perimenopause and with or without menorrhagia. We hypothesized that differences in vessel numbers and types exhibiting alpha-actin staining would exist between these groups, reflecting structural/functional differences. The results showed that perimenopausal menorrhagic women had significantly more smooth muscle alpha-actin expression than non-perimenopausal controls in four out of five menstrual cycle stages (P < 0.05), while perimenopausal non-menorrhagic women demonstrated a significant increase at the mid-proliferative stage only (P < 0.007). No significant differences occurred between women with or without menorrhagia before or during the perimenopause. Perimenopausal women had significantly more straight arterioles (P < 0.02) than women prior to perimenopause at the late secretory stage, while non-perimenopausal women demonstrated significantly higher numbers of spiral arterioles (P < 0.002) in the early secretory stage, although this difference had disappeared by the late secretory stage. In conclusion, we found no major differences in endometrial vascular smooth muscle alpha-actin staining between women with and without menorrhagia, but significant increases in alpha-actin staining in women showing perimenopausal symptoms.

Production of an endothelial cell migratory signal in rat endometrium during early pregnancy.

Rat endometrial explants were cultured in a three-dimensional collagen/endothelial cell matrix to measure angiogenic activity, as represented by migration of vascular endothelial cells towards the explants. Minimal endothelial cell migratory activity was observed with endometrial explants taken during the four-day oestrous cycle and days 3 and 4 of pregnancy. In contrast, a surge of endothelial cell migration occurred in response to endometrial explants taken from day-5-pregnant rats. Activity was found in explants taken approximately 5 h prior to implantation, but returned to minimal levels by day 6 of pregnancy. Endothelial cell migration remained minimal in response to both implantation and intersite tissue explants taken from days 6 and 7 of pregnancy. Endometrium from ovariectomised rats produced no endothelial cell migratory activity as measured by this technique. However, near preimplantation levels of endothelial cell migratory activity could be induced in ovariectomised rat endometrium by administering progesterone for 72 hours. Oestrogen given in conjunction with progesterone had no additional effect. These results demonstrate the presence of an endometrial signal that controls endothelial cell migration, and demonstrate this activity can be induced by progesterone without the addition of oestrogen.

Human follicular fluid maturity and endothelial cell chemotaxis.

Follicular fluid of varying maturity was collected from the largest Graafian follicle of 23 ovulatory patients during laparoscopy (five blood-stained samples were discarded) and from five patients undergoing oocyte collection for assisted reproduction. The endothelial cell (EC) chemotactic potentials of the samples were tested with bovine aortic ECs in modified Boyden Chambers and an EC density score was determined for each sample. Intra-assay coefficients of variation (CV) varied from 2.9-17%; inter-assay CV was 45%. Therefore, cell density scores were expressed in terms of positive and negative control values from their respective plates. Serum progesterone was positively correlated with EC chemotactic potential (R2 = 14.9%). There was no correlation with day of cycle, follicular diameter, serum luteinizing hormone (LH) or oestradiol, or follicular fluid total protein levels (all R2 < or = 6%). Five subjects were in early and four in mid-follicular phase, seven were in early and two in late LH surge, and there were also five in-vitro fertilization (IVF) specimens. There was a statistically significant difference between the EC chemotactic potential of the five groups by analysis of variance (F = 5.98; P = 0.006). There was no significant difference between the late LH surge and the IVF samples (two-sample t-test). The mean cell density was higher for these two groups than the other three (P < 0.01 in all cases; two-sample t-test). It is concluded that the EC chemotactic potential of human follicular fluid increases after the LH surge and before ovulation.

Endometrial sex steroid receptor expression in women with menorrhagia.

OBJECTIVE: To investigate the oestrogen and progesterone receptor distribution in endometrium from reproductive age and perimenopausal women with and without menorrhagia. DESIGN: A comparative observational study. SUBJECTS: Forty-five women with objective menorrhagia (27 categorised as reproductive age and 18 as perimenopausal) and a control group of 44 women (31 reproductive age and 13 perimenopausal) with menstrual blood loss of less than 80 ml per period. MAIN OUTCOME MEASURES: Oestrogen receptor and progesterone receptor semi-quantitative immunostaining scores. RESULTS: Comparison of control and menorrhagic endometrium in this study (whether from reproductive age or perimenopausal subjects) failed to demonstrate any major differences in either sex steroid receptor mean immunostaining score. The results demonstrated a great degree of variability in sex steroid receptor immunoreactivity between individuals. Irrespective of clinical group, significant increases in immunostaining were demonstrated in the proliferative phase of the cycle for immunoreactivity of oestrogen receptor in glands (P < 0.0005), and stromal (P = 0.002) compartments of endometrium and progesterone receptor immunoreactivity in glands (P = 0.009). Progesterone receptor immunostaining in the stromal compartments did not significantly decline (P = 0.06) in the secretory phase. CONCLUSIONS: Endometrium from women with objective evidence of menorrhagia is indistinguishable in terms of sex steroid immunoreactivity from endometrium of women with normal monthly blood loss. This pattern of sex steroid receptor immunostaining pattern was maintained into the perimenopausal years.

Human follicular fluid maturity and endothelial cell mitogenesis.

Follicular fluid, of varying maturity, (day 5-16 of cycle) was collected from the largest Graafian follicle of each of 22 ovulatory patients during laparoscopic procedures. Three samples were blood-stained and discarded. The mitogenic potential of each sample was determined using bovine aortic endothelial cells in the CellTiter 96 Non-Radioactive Cell Proliferation/Cytotoxicity Assay system. Intra- and inter-plate coefficients of variation were < 9%. The follicular fluid samples induced cell doubling times which varied from approximately 12-24 h and final cell numbers which, in the individual wells, ranged from 7828-30,900 (starting number 2000/well). Follicular fluid total protein content was unrelated to the mitogenic potential, (R2 = 0%). Serum oestradiol was negatively correlated with the mitogenic potential (R2 = 26%). No correlation was found with day of the menstrual cycle (R2 = 4.3%), maximum follicular diameter (R2 = 1.8%), or serum concentration of progesterone (R2 = 0.7%), luteinizing hormone (LH) (R2 = 1.5%) or follicle stimulating hormone (R2 = 0.1%). Five subjects were in 'early' and six in 'mid'-follicular phase, six were in 'early' and two in 'late' LH surge. There was no difference in the mitogenic response between these four groups by one-way analysis of variance (F = 0.21; P = 0.89). It is concluded that the mitogenic potential of human follicular fluid is not related to Graafian follicle maturity or, more particularly, to the LH surge.

Endothelial cell migratory signal produced by human endometrium during the menstrual cycle.

Human endometrial explants were cultured in a three-dimensional collagen/bovine aortic endothelial cell (BAEC) matrix to measure angiogenic activity, as represented by migration of BAEC towards the explants. The 57 endometrial biopsies were classified by histological appearance into nine stages of the menstrual cycle; five proliferative groupings, three secretory groupings and one menstrual. The BAEC migratory score was taken as the average for 12 explants assayed from each biopsy. The results showed two significant peaks of BAEC migratory activity, one during the early proliferative phase and one during the mid--late proliferative phase. There was a significant drop in the BAEC migratory signal from early--mid-proliferative endometrial explants compared to most other stages of the cycle. The results also show a non-significant rise in BAEC migratory activity in the mid-secretory phase of the cycle. Overall, the results support the concept of two or three different endometrial angiogenic events during the human menstrual cycle, a post-menstrual repair, a mid--late proliferative growth and a lesser mid-secretory activity that may be associated with spiral arteriole growth. Each of these events occurs under different hormonal environments and will need to be investigated separately in terms of local mechanisms controlling angiogenesis.