To study the role of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF-2) in the development of sensory neurons, the cochleovestibular ganglion of the chicken embryo provides a well-characterized structure. This permits use of morphological markers in a cell culture preparation comparable to the normal embryo (normocytic). Otocysts were explanted from white leghorn embryos at Hamburger-Hamilton Stages 14-16, when ganglion cell precursors normally start migrating from the otic epithelium. The cultures were supplemented with either fetal bovine serum or human recombinant FGF-2 (in defined medium or serum) for 2 or 5 days. FGF-2 increased explant <em>growth</em>, neuroblast migration, and neurite out<em>growth</em> 2- to 10-fold in the first 2 days. Neuronal morphology appeared within 2-3 days with FGF-2 but required at least 4-5 days with serum. FGF-2 in defined medium stimulated early migration and differentiation, but without serum led to degeneration after 5 days. In serum, <em>growth</em> was later and slower but continued for at least 3 weeks. When explants were cultured in serum with a neutralizing antibody to FGF-2, but no FGF added, neuroblast migration and elongation were decreased by 2- to 4-fold, compared to serum alone. Immunocytochemistry demonstrated FGF receptor sites on the migrating ganglionic neuroblasts, on their processes and <em>growth</em> cones, and in the incipient ganglion and otic epithelium at Stages 15-<em>17</em>, both in the embryo and in vitro. The findings suggest that FGF-2 stimulates early migration and differentiation of ganglion cells by activating the receptors of neuroblasts or their precursors in the embryonic otocyst. However, other <em>factors</em> must sustain their later development.

The biological effects of different wavelengths of light emitting diode (LED) light tend to vary from each other. Research into use of photobiomodulation for treatment of skin wounds and the underlying mechanisms has been largely lacking. We explored the histopathological basis of the therapeutic effect of photobiomodulation and the relation between duration of exposure and photobiomodulation effect of different wavelengths of LED in a Japanese big-ear white rabbit skin-wound model. Skin wound model was established in 16 rabbits (three wounds per rabbit: one served as control, the other two wounds were irradiated by red and blue LED lights, respectively). Rabbits were then divided into 2 equal groups based on the duration of exposure to LED lights (15 and 30 min/exposure). The number of wounds that showed healing and the percentage of healed wound area were recorded. Histopathological examination and skin expression levels of <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF), epidermal <em>growth</em> <em>factor</em> (EGF), endothelial marker (CD31), proliferating cell nuclear antigen (Ki67) and macrophagocyte (CD68) infiltration, and the proliferation of skin collagen fibers was assessed. On days 16 and <em>17</em> of irradiation, the healing rates in red (15 min and 30 min) and blue (15 min and 30 min) groups were 50%, 37.5%, 25% and 37.5%, respectively, while the healing rate in the control group was 12.5%. The percentage healed area in the red light groups was significantly higher than those in other groups. Collagen fiber and skin thickness were significantly increased in both red light groups; expression of EGF, FGF, CD31 and Ki67 in the red light groups was significantly higher than those in other groups; the expression of FGF in red (30 min) group was not significantly different from that in the blue light and control groups. The effect of blue light on wound healing was poorer than that of red light. Red light appeared to hasten wound healing by promoting fibrous tissue, epidermal and endothelial cell proliferation. An increase in the exposure time to 30 min did not confer any additional benefit in both red and blue light groups. This study provides a theoretical basis for the potential therapeutic application of LED light in clinical settings.

BACKGROUND

Focus on non-classical effects and possible less side effects of treatment with nutritional vitamin D, raises the expectation of possible benefits from treating chronic kidney disease (CKD) patients with ergocalciferol (vitamin D2). Treatment with 1,25(OH)2 vitamin D (calcitriol) induces elevated fibroblast growth factor 23 (FGF23), while epidemiological studies have found positive effects of nutritional and 25(OH)vitamin D on mortality in CKD. Disturbed mineral homeostasis in CKD is correlated to adverse outcome and cardiovascular mortality. The objective was to examine the possible effects of treatment with high doses of ergocalciferol on parameters of mineral homeostasis in predialysis CKD patients.

METHODS

A total of 43 adult patients with CKD stage 4-5, not receiving vitamin D supplementation, were studied, and allocated by simple randomization to either an intervention (n = 26) or a control group (n = 17). The intervention group received ergocalciferol, 50.000 IU/week for 6 weeks. Plasma FGF23, creatinine, parathyroid hormone (PTH), phosphate and ionized calcium were obtained at baseline and after the 6 weeks.

RESULTS

The intervention group had a significant increase in 25(OH)D2 concentration from < 10 to 90 ± 4 nmol/L, while 1,25(OH)2D (62 ± 6 at baseline and 67 ± 6 pmol/L at 6 weeks) remained stable. No changes were seen in the circulating vitamin D concentrations in the control group. After the 6 weeks of treatment no significant changes were seen in concentration of creatinine, phosphate, ionized calcium, PTH and FGF23 remained stable.

CONCLUSIONS

No harmful effects of short-term treatment with high-dose ergocalciferol were observed on markers of mineral homeostasis and FGF23 in CKD patients stage 4-5.

Cancer cachexia is a multifactorial syndrome that dramatically decreases survival. Loss of white adipose tissue (WAT) is one of the key characteristics of cachexia. WAT wasting is paralleled by microarchitectural remodeling in cachectic cancer patients. Fibrosis results from uncontrolled ECM synthesis, a process in which, transforming growth factor-beta (TGFβ) plays a pivotal role. So far, the mechanisms involved in adipose tissue (AT) re-arrangement, and the role of TGFβ in inducing AT remodeling in weight-losing cancer patients are poorly understood. This study examined the modulation of ECM components mediated by TGFβ pathway in fibrotic AT obtained from cachectic gastrointestinal cancer patients.

After signing the informed consent form, patients were enrolled into the following groups: cancer cachexia (CC, n = 21), weight-stable cancer (WSC, n = 17), and control (n = 21). The total amount of collagen and elastic fibers in the subcutaneous AT was assessed by histological analysis and by immunohistochemistry. TGFβ isoforms expression was analyzed by Multiplex assay and by immunohistochemistry. Alpha-smooth muscle actin (αSMA), fibroblast-specific protein (FSP1), Smad3 and 4 were quantified by qPCR and/or by immunohistochemistry. Interleukin (IL) 2, IL5, IL8, IL13 and IL17 content, cytokines known to be associated with fibrosis, was measured by Multiplex assay.

There was an accumulation of collagen and elastic fibers in the AT of CC, as compared with WSC and controls. Collagens type I, III, VI, and fibronectin expression was enhanced in the tissue of CC, compared with both WSC and control. The pronounced expression of αSMA in the surrounding of adipocytes, and the increased mRNA content for FSP1 (20-fold) indicate the presence of activated myofibroblasts; particularly in CC. TGFβ1 and TGFβ3 levels were up-regulated by cachexia in AT, as well in the isolated adipocytes. Smad3 and Smad4 labeling was found to be more evident in the fibrotic areas of CC adipose tissue.

Cancer cachexia promotes the development of AT fibrosis, in association with altered TGFβ signaling, compromising AT organization and function.

Elevated levels of <em>fibroblast</em> <em>growth</em> <em>factor</em> 23 (FGF23) and phosphorus (P) have been linked to greater risks of left ventricular hypertrophy (LVH) in patients with end stage renal disease (ESRD). The aim of this study was to test if differences exist in a long nocturnal HD group in comparison with a group treated with standard daily thrice weekly dialysis. The attempt was to evaluate if elevated FGF-23 levels, intact parathyroid hormone and P might be associated with left ventricular mass index (LVMI). Quantitative echocardiographic analyses were performed at baseline in 50 maintenance HD patients (<em>17</em> women and 33 men, mean age: 56.4 ± 15.35 years, mean HD vintage: 9.06 ± 8.86 years, all patients are on HD thrice a week-median duration 15 h/week, 10 of them on long nocturnal HD, median duration 24 h/week). LVMIs were calculated. FGF23 was measured in duplicate using a second generation C-terminal enzyme-linked immunosorbent assay and log of FGF-23 values were computed. Mean LVMI was 136.44 ± 44.44 g/m(2) . Serum FGF-23 levels were elevated when compared to population data with preserved kidney function (median 1388.5 RU/mL, range 252 to 24 336 RU/mL). There were no correlations recorded between log FGF-23 levels and LVMI (r = 0.2, P = 0.66). LVMI was significantly lower in HD patients on long nocturnal dialysis procedure (r = -0.31, P = 0.05). Patients treated with long nocturnal HD showed lower LV mass, lower P-values and higher 25-OH-D3 supply. Plasma FGF-23 concentration was comparable between the groups and was not associated with LVMI in our maintenance HD patients.

During development, Purkinje axons elongate along precise trajectories and acquire stereotypic branching patterns to innervate targets in the deep nuclei and cerebellar cortex. These processes are accomplished through cell-intrinsic mechanisms, whose operation is regulated by environmental signaling cues. Here, we show that Anosmin-1, the protein defective in the X-linked form of Kallmann syndrome, is one among such cues. Anosmin-1, that stimulates axon elongation and branching in the ol<em>factor</em>y system, is expressed by Purkinje cells and deep nuclear neurons of the rat cerebellum during the ontogenetic period when Purkinje axons acquire their mature pattern. These neurons also express the putative Anosmin-1 receptor, <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor 1. Application of Anosmin-1 to dissociated cultures of embryonic (embryonic day <em>17</em>, E<em>17</em>) or postnatal (postnatal day 0, P0) rat cerebellar cells enhances neuritic elongation and exerts a strong promoting action on the budding of collateral branches and on the extension of terminal arbors. Opposite effects are observed when neutralizing anti-Anosmin-1 antibodies are applied to the same cultures. Comparable results are obtained by administering the protein or the blocking antibodies to organotypic cultures of postnatal (P0) rat cerebellum. In P10 cerebellar slices, Anosmin-1 does not enhance the spontaneous regenerative capabilities of severed Purkinje axons, but promotes the terminal out<em>growth</em> of injured neurites into embryonic neocortical explants apposed to the axotomy site. Although Anosmin-1 is unable to change the overall intrinsic <em>growth</em> competence of Purkinje cells, it exerts a powerful stimulatory action on the budding and extension of collateral branches and terminal plexus, contributing to the patterning of Purkinje axons.

The potential role of satellite cells in mediating the effect of trenbolone [<em>17</em> beta-hydroxyestra-4,9-11-trien-3-one (TBOH)] on skeletal muscle hypertrophy was examined. Young female Sprague-Dawley rats received TBOH injections daily for 2 weeks; <em>growth</em>, body composition, and the composition of selected muscles were assessed. Treated rats grew more rapidly and deposited less body lipid and more protein. The semimembranosus muscle from treated rats was larger and had approximately 60% more DNA per muscle than muscles from control rats. The addition of trenbolone directly to the medium of cultured satellite cells did not stimulate cell proliferation, nor did it augment the stimulatory response of these cells to <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) or insulin-like <em>growth</em> <em>factor</em> I (IGF-I). In contrast, satellite cells cultured from TBOH-treated rats exhibited greater proliferative responses to FGF and IGF-I than satellite cells from control rats. In addition, serum from TBOH-treated rats stimulated greater cell proliferation in satellite cell cultures than serum from control rats. These experiments suggest that one possible mechanism responsible for the ability of TBOH to stimulate skeletal muscle hypertrophy may be through enhanced proliferation and differentiation of satellite cells as a result of the increased sensitivity of these cells to IGF-I and FGF.

OBJECTIVE

We examined the safety and efficacy of controlled-release basic fibroblast growth factor (b-FGF) for peripheral artery disease (PAD), compared with autologous bone marrow mononuclear cell implantation (BMCI).

BACKGROUND

We recently developed a b-FGF-incorporated biodegradable hydrogel that enables slow-releasing drug delivery system.

METHODS

PAD patients were divided into a b-FGF group (n=10) and BMCI group (n=15). Injection of gelatin hydrogel containing 600 μg b-FGF or BMCI (0.4-5.1×10(10) cell) was performed. Visual analog pain scale (VAS), (99m)technetium-tetrofosmin (Tc-TF) scintigraphy, transcutaneous oxygen tension (TcPO(2)), and ankle-brachial index (ABI) were evaluated before and 4 weeks after each treatment, and 2-year prognosis was determined.

RESULTS

VAS (b-FGF 67±15 to 4±5, p<0.01, BMCI 67±42 to 5±9 mm, p<0.01) and TcPO(2) (b-FGF 16±14 to 47±17, p<0.01, BMCI 13±13 to 37±21 mmHg, p<0.01) were significantly improved in both groups. Tc-TF and ABI were not changed. Prognosis was similar between the groups (b-FGF 91%, BMCI 80%, NS).

CONCLUSIONS

Controlled-release b-FGF is as safe as BMCI, and its efficacy appears to be comparable. Thus, this therapy may be an alternative to BMCI.

The <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) family comprises to date 12 members, which are involved in various physiological processes throughout embryogenesis and adult life. Two novel members of the family have been identified recently (FGF11 and FGF12). Using in situ hybridization on metaphasic chromosomes, we have been able to assign FGF11 to band p12-p13 of human chromosome <em>17</em> and FGF12 to band q28 of human chromosome 3.

BACKGROUND

Risk factors for diastolic dysfunction in hypertrophic cardiomyopathy (HCM) are poorly understood. We investigated the association of variants in hypoxia-response genes with phenotype severity in pediatric HCM.

METHODS

A total of 80 unrelated patients <21 y and 14 related members from eight families with HCM were genotyped for six variants associated with vascular endothelial growth factor A (VEGFA) downregulation, or hypoxia-inducible factor A (HIF1A) upregulation. Associations between risk genotypes and left-ventricular (LV) hypertrophy, LV dysfunction, and freedom from myectomy were assessed. Tissue expression was measured in myocardial samples from 17 patients with HCM and 20 patients without HCM.

RESULTS

Age at enrollment was 9 ± 5 y (follow-up, 3.1 ± 3.6 y). Risk allele frequency was 67% VEGFA and 92% HIF1A. Risk genotypes were associated with younger age at diagnosis (P < 0.001), septal hypertrophy (P < 0.01), prolonged E-wave deceleration time (EWDT) (P < 0.0001) and isovolumic relaxation time (IVRT) (P < 0.0001), and lower freedom from myectomy (P < 0.05). These associations were seen in sporadic and familial HCM independent of the disease-causing mutation. Risk genotypes were associated with higher myocardial HIF1A and transforming growth factor B1 (TGFB1) expression and increased endothelial-fibroblast transformation (P < 0.05).

CONCLUSIONS

HIF1A-upregulation and/or VEGFA-downregulation genotypes were associated with more severe septal hypertrophy and diastolic dysfunction and may provide genetic markers to improve risk prediction in HCM.

We established an in vitro hypoxia model and investigated the protective effect of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) against neuronal cell death caused by hypoxia. Hippocampal neurons obtained from rats on embryonic day (E) <em>17</em> and 20 and on postnatal day (P) 4 were cultured for 6-24 h in an oxygen-deprived state. This in vitro hypoxia study showed that the cultured neurons were sensitive to the oxygen deprivation. The cultured P4 rat hippocampal neurons seemed to be weaker in the hypoxia condition than those of E<em>17</em> and E20 rats, suggesting that the cultured postnatal cells might be sensitive to hypoxia. bFGF, but not nerve <em>growth</em> <em>factor</em>, prevented the neuronal cell death caused by hypoxia in a dose-dependent manner.

Since the cornea is an avascular tissue, the wound healing process is lengthy, with a need for sutures to stabilize the wound for a long time. Platelet-derived <em>growth</em> <em>factor</em> (PDGF) has been shown to accelerate wound healing in rat dermal models. Accelerated healing, if unaccompanied by side effects may reduce suture related complications such as astigmatism and infectious keratitis. This study evaluated the effect of PDGF on wound strength in corneal laceration and penetrating keratoplasty models using New Zealand white albino rabbits. Twenty-two rabbits were used in the corneal laceration model and sixteen rabbits in the penetrating keratoplasty model. The treated rabbits received 385 picomoles/drop of PDGF-BB dissolved in balanced salt solution six times on day 1 and three times a day for the remainder of the study. The control rabbits received balanced salt solution in the same dosing schedule. The pressure required to rupture the wound was measured using a pressure transducer. In the laceration model the PDGF treated group had mean (+/- standard deviation) average pressures on day 7 of 360 +/- 102 mm Hg for wound rupture compared to 210 +/- 102 mm Hg in the control group. (p = 0.005). The average pressures in the penetrating keratoplasty model on day <em>17</em> were 707 +/- 201 mm Hg for the controls and 1042 +/- 292 mm Hg for the PDGF treated group (p = 0.026). Histopathological evaluation of eyes not subjected to bursting showed increased <em>fibroblasts</em> at the wound junction with an increase in types III and type IV collagen production.(ABSTRACT TRUNCATED AT 250 WORDS)

Transforming <em>Growth</em> <em>Factor</em>-beta (TGF-beta) inhibits the serum and basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF)-induced proliferation of cultured bovine retinal endothelial capillary (BREC) cells in a dose-dependent manner. The concentration of TGF-beta required to get half-maximal inhibition (ED50) are 10 pg ml-1 in serum and <em>17</em> pg ml-1 in the presence of additional bFGF (1 ng ml-1). These TGF-beta ED50 values are greatly increased when BREC cells were seeded at high density: 610 pg ml-1 in serum and 1 ng ml-1 in the presence of additional bFGF. At low initial cell density BREC cells are more sensitive to TGF-beta than aortic bovine arch endothelial (ABAE) cells for which TGF-beta ED50 values are respectively 40 pg ml-1 and 100 pg ml-1 in serum and in the presence of additional bFGF. In contrast, at high cell density BREC cells appeared to be more resistant to TGF-beta inhibition than ABAE cells for which TFG-beta ED50 values are 210 and 300 pg ml-1. Moreover bFGF added at increasing concentrations neutralize totally TGF-beta inhibition of BREC cell proliferation but only partially that of ABAE cell proliferation. Our results suggest a key role of equilibrium TFG-beta bFGF on the proliferation of BREC cells.

Flexor tendon wound healing in zone II is complicated by adhesions to the surrounding fibro-osseous sheath. These adhesions can significantly alter tendon gliding and ultimately hand function. Lactate and transforming <em>growth</em> <em>factor</em>-beta (TGF-beta) are two important mediators of wound healing that have been demonstrated to independently increase collagen production by cells of the tendon sheath, epitenon, and endotenon. This study examined the effects of lactate on TGF-beta peptide and receptor production by flexor tendon cells. Tendon sheath <em>fibroblasts</em>, epitenon tenocytes, and endotenon tenocytes were isolated from rabbit flexor tendons and cultured separately. Cell cultures were supplemented with 50 mM lactate, and the expression of three TGF-beta peptide isoforms (beta1, beta2, and beta3) and three receptor isoforms (R1, R2, and R3) was quantified with enzyme-linked immunosorbent assays. TGF-beta functional activity was also assessed with the addition of tendon cell conditioned media to mink lung epithelial cells transfected with a luciferase reporter gene expression construct responsive to TGF-beta. Supplementation of the cell culture medium with lactate significantly (p < 0.05) increased the expression of all TGF-beta peptide and receptor isoforms in all three cell lines. Tendon sheath <em>fibroblasts</em> exhibited the greatest increases in beta1 and beta2 peptide isoform expression (30 and 23 percent, respectively), whereas endotenon tenocytes demonstrated the greatest increase in beta3 peptide expression (32 percent). Epitenon tenocytes exhibited the greatest increases in receptor isoform R1 and R2 expression (<em>17</em> and 19 percent, respectively). All three tendon cell types demonstrated significant (p < 0.05) increases in TGF-beta functional activity when exposed to lactate. Epitenon tenocytes demonstrated the greatest increase in activity (>4 times control values), whereas tendon sheath <em>fibroblasts</em> demonstrated the highest overall levels of total TGF-beta functional activity. Lactate significantly increased TGF-beta peptide (beta1, beta2, and beta3) expression, receptor (R1, R2, and R3) expression, and functional activity, suggesting a common pathway regulating tendon cell collagen production. Modulation of lactate and TGF-beta levels may provide a means of modulating the effects of TGF-beta on adhesion formation in flexor tendon wound healing.

Peripheral blood cells (PBCs) collected by leukapheresis after progenitor mobilization with chemotherapy and <em>growth</em> <em>factors</em> have been used successfully to replace marrow autografts in protocols requiring stem-cell support. Moreover, such transplants are often associated with more rapid recovery of blood cell counts than is routinely achieved with bone marrow. While conditions that mobilize colony-forming cells (CFCs) into the circulation are becoming increasingly well characterized, little information is available as to how these or other mobilizing treatments may influence the release of more primitive cells into the peripheral blood. To quantitate the peripheral blood content of such cells, we used the long-term culture-initiating cell (LTC-IC) assay, which detects a cell type that is able to produce progeny CFCs after a minimum of 5 weeks in cultures containing marrow <em>fibroblasts</em>. In this report, we present the findings on 21 patients who were transplanted over a 7-year period at our institution with PBCs alone. PBCs were collected in steady-state (n = 6) or during the recovery phase after high-dose cyclophosphamide (Cy; n = 15, nine with and six without additional <em>growth</em> <em>factor</em> administration). PBCs collected from another 11 patients given granulocyte colony-stimulating <em>factor</em> (G-CSF) were transplanted together with autologous marrow. Time-course studies of nine patients after Cy +/- granulocyte-macrophage CSF (GM-CSF) showed that CD34+ cells, CFCs, and LTC-ICs fell from normal to undetectable levels after Cy, and increased at the time of white blood cell (WBC) recovery: LTC-ICs to a mean of sixfold and CFCs to a mean of 26-fold higher than normal. The mean number of CD34+ cells, CFCs, and LTC-ICs present in the PBC harvest was twofold to 10-fold higher after mobilization than in steady-state collections; however, more than 2-log interpatient variability was observed. After PBC transplantation, the median time to a WBC count more than 10(9)/L was 12 days; polymorphonuclear leukocyte (PMN) count more than 0.5 x 10(9)/L, 15 days; and platelet count more than 20 x 10(9)/L, <em>17</em> days, although patients who received fewer than 1.5 x 10(5) CFCs/kg had a more than 50% chance of delayed count recovery >> 28 days). Patients who received Cy + GM-CSF-stimulated PBCs had more rapid and consistent platelet recoveries as compared with other groups receiving Cy mobilized or steady-state PBCs alone, and a rapid WBC recovery after Cy predicted a rapid WBC recovery after transplantation.(ABSTRACT TRUNCATED AT 400 WORDS)

<em>Fibroblast</em> <em>growth</em> <em>factors</em> (FGFs) are secreted regulatory proteins involved in various developmental processes. In vertebrates, the FGF superfamily comprises 22 members. In non-vertebrates, six FGF genes have been identified in Ciona intestinalis, three in Drosophila melanogaster, and two (let-756 and egl-<em>17</em>) in Caenorhabditis elegans. The core of LET-756 shares a 30-50% sequence identity with the various members of the superfamily. The relationships between vertebrate and non-vertebrate FGFs are not clear. We made chimeric FGFs by replacing the core region of LET-756 by the cores of various mammalian, fly, and worm FGFs. LET-756 deleted in its core region was no longer able to rescue the lethal phenotype of a let-756 null mutant, and only chimeras containing the cores of FGFs 9, 16, and 20 showed rescue capacity. This core contains an internal motif of six amino acid residues (EFISIA) whose deletion or mutation abolished both the rescue activity and FGF secretion in the supernatant of transfected COS-1 cells. Chimera containing the core of C. intestinalis FGF9/16/20, a potential ortholog of FGF9 lacking the complete EFISIA motif, was not able to rescue the lethal phenotype or be secreted. However, the introduction of the EFISIA motif restored both activities. The data show that the EFISIA motif in the core of LET-756 is essential for its biological activity and that FGFs 9, 16, and 20, which contain that motif, are functionally close to LET-756 and may be evolutionary related. This non-classical mode of secretion using an internal motif is conserved throughout evolution.

BACKGROUND

Trophic factors [e.g. basic fibroblast growth factor (bFGF)] released by transplanted retinal pigment epithelial (RPE) cells are able to slow down the hereditary degeneration of the retina in the Royal College of Surgeons rat in sites distant from the site of transplantation where rod outer segment (ROS) phagocytic activity is not reconstituted by the transplants.

METHODS

To investigate whether iris pigmented epithelial (IPE) cells are also able to generate this rescue by trophic factors, we transplanted IPE cells from Long-Evans rats into the choroid and subretinal space of 17 young RCS rats. The eyes were enucleated after 6 months and prepared for light microscopy. Six age-matched RCS rats served as controls. Light microscope sections from the whole choroid, healthy choriocapillaris, transplanted cells and the maximum thickness of the choroid, and outer nuclear layer parameters were analyzed by computer-assisted morphometry.

RESULTS

In transplanted animals photoreceptor cells were rescued from degeneration although the majority of the transplanted IPE cells were located in the choroid. In the non-transplanted group photoreceptors were absent.

CONCLUSIONS

Transplantation of IPE cells slows down degeneration of the photoreceptors in the RCS rat. This photoreceptor-sparing effect by the IPE cells was observed even when the transplants were predominantly located within the choroid. The beneficial effect observed may be related to trophic factors possibly secreted by the transplanted IPE cells.

BACKGROUND

Various pathogenetic mechanisms have been proposed to explain the development of antrochoanal polyps (ACPs); however, the cause is still largely unknown. The aim of this study was to characterize the expression and the potential role of a battery of molecular markers in the development of ACPs. A prospective controlled study of a case series was performed.

METHODS

Tissue samples of maxillary sinus mucosa were obtained from 14 patients with ACPs, <em>17</em> patients with chronic nonpolypoid maxillary sinusitis, and 4 patients with normal maxillary sinus mucosa; RNAs were extracted from the sinus mucosa, and semiquantitative reverse transcription-polymerase chain reaction was performed for basic <em>fibroblast</em> <em>growth</em> <em>factor</em>, transforming <em>growth</em> <em>factor</em> P, and mucin genes (MUC), MUC5AC, MUC5B, and MUC8, to investigate their expression.

RESULTS

The expression of basic fibroblast growth factor and transforming growth factor beta was significantly higher in ACPs than in chronic rhinosinusitis and healthy mucosa. Meanwhile, the levels of expression of MUC genes were higher in ACPs and chronic rhinosinusitis compared with healthy mucosa.

CONCLUSIONS

These findings suggest that ACPs may represent an inflammatory reaction caused by overproduction of tissue-derived growth factors in an inductive environment.

The effects of medroxyprogesterone acetate (MPA) (I) and related compounds (II-VI) upon angiogenesis induced by basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) or transforming <em>growth</em> <em>factor</em>-alpha (TGF-alpha) were investigated using a rabbit corneal system for assay of angiogenesis. Dexamethasone (Dex) was used as a positive control. The MPA analogues tested were 6,6'-dehydro-MPA (II), megestrol acetate (III), 1-dehydromegestrol acetate (IV), melengestrol acetate (V), and 1-dehydromelengestrol acetate (VI). The inhibitory activities of these steroids using bFGF were in the order: Dex = MPA = (VI) = (V)>> (IV)>> (III). Steroid (II) was inactive. 5 alpha-dihydrotestosterone was weakly active, while estradiol-<em>17</em> beta and progesterone were inactive. The angiostatic activity of MPA was completely abolished by mefipristone (RU 486) which showed no anti-angiogenic activity in this assay. With TGF-alpha, the order of angiostatic activities was Dex = (VI)>> (IV)>> (III)>> (V). Steroid (II) was again inactive. Dex, MPA, and all the MPA analogues except steroid (II) markedly inhibited the activity of plasminogen activator secreted by cultured calf pulmonary artery endothelial cells, but did not inhibit <em>growth</em> of these cells. The binding affinities of MPA and its analogues to glucocorticoid, progesterone and androgen receptors were determined, but were found not to be correlated with their angiostatic activities.

We have previously shown that inhibition of nitric oxide generated by inducible nitric oxide synthase (iNOS) results in impaired colon anastomotic healing. Therefore, we proceeded to assess whether disruption of iNOS activity alters the normal pattern of <em>growth</em> <em>factor</em> expression during anastomotic healing. Two groups of male Sprague-Dawley rats underwent distal colonic division and anastomosis, jugular venous catheterization and subcutaneous placement of polyvinyl alcohol sponges. The first group (n = 10) received q8 hour intravenous injections of 10 mg/kg L-N-iminoethyl-lysine (L-NIL, a selective inhibitor of iNOS), while the second group (n = 12) received equal volumes of saline. On postoperative day 5, animals were sacrificed and anastomotic bursting pressure was determined. Histologic sections of the anastomosis were subjected to in situ hybridization versus mRNA of the proteins listed below. Positive controls were reacted with a poly-thymidine (poly-T) probe versus ubiquitous mRNA poly-adenine tails. Positively stained cells were quantified using a calibrated optical grid encompassing 0.5 mm(2) area centered over the anastomosis. Results are reported as the number of positive cells per 1000 cells positive for poly-T. L-NIL treated animals demonstrated an 18% decrease in wound fluid NO(X) compared to controls (29.2 +/- 1.2 vs. 34.6 +/- 2.0 microM, mean +/- SEM; P = 0.035). This corresponded to a <em>17</em>% decrease in anastomotic bursting pressure (153 +/- 4 vs. 182 +/- 8 mm Hg, mean +/- SEM; P < 0.05). L-NIL also markedly increased the number of cells expressing transforming <em>growth</em> <em>factor</em>-beta, tumor necrosis <em>factor</em>-alpha, vascular endothelial <em>growth</em> <em>factor</em>, and both inducible and endothelial forms of nitric oxide synthase. L-NIL had no effect on the expression of basic <em>fibroblast</em> <em>growth</em> <em>factor</em>. The data demonstrate that iNOS inhibition markedly disrupts the profile of cytokine and <em>growth</em> <em>factor</em> mRNA normally expressed during anastomotic healing. This provides in vivo evidence that NO modulates gene expression during anastomotic healing.

In a previous study, we found that basic <em>fibroblast</em> <em>growth</em> <em>factor</em> could stimulate bone-graft incorporation. In the present study, the effects of different doses and implantation times were further studied, using the bone conduction chamber, in rats. Inside the chamber, the graft is isolated from the surrounding tissues except at one end, where small openings embedded in host bone allow in<em>growth</em> of tissue. The distance that new tissues had reached from the openings into the graft was measured on histological slides. Bone grafts were obtained from the proximal tibiae of donor rats, frozen at -70 degrees C, and lipid-extracted. Before implantation, they were soaked overnight in a hyaluronate gel with or without basic <em>fibroblast</em> <em>growth</em> <em>factor</em> and then were fitted into the chambers, which were implanted in the proximal tibiae of recipient rats. In a dose-response experiment, grafts containing 0.3, 8, 40, 200, or 1,000 ng of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> were compared with grafts treated with carrier gel only, after an implantation time of 6 weeks. Fibrous tissue always penetrated the grafts further than the ingrown bone; the distance that it reached from the in<em>growth</em> openings (total in<em>growth</em> distance) was increased by all of the doses except 0.3 ng per implant. The distance of bone in<em>growth</em> was increased by 8, 40, and 200 ng. The increased total in<em>growth</em> with 1,000 ng was due to an increased amount of fibrous tissue ahead of the bone, whereas with the lower doses the increase was due to more bone. Thus, the dose had an effect on the type of ingrown tissue found in the graft. In a time-effect study, grafts treated with 40 ng of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> had a higher uptake of [99mTc]MDP at 2 and 4 weeks and an increased bone in<em>growth</em> distance at 10 weeks. The radioactivity from [125I]basic <em>fibroblast</em> <em>growth</em> <em>factor</em> declined with a half-life of <em>17</em> hours. The results suggest that basic <em>fibroblast</em> <em>growth</em> <em>factor</em> may be beneficial for the incorporation of contained bone grafts; studies using more clinically relevant models are required.

OBJECTIVE

This study examined the potential of amniotic fluid mesenchymal stem cells (AF-MSCs) to generate lung precursor cells in vitro and on a xenologous three-dimensional de-cellularized lung scaffold.

METHODS

AF-MSCs were isolated from human amniotic fluid obtained from <em>17</em>-37 weeks gestation. Lung differentiation was induced on Matrigel or on de-cellularized rat lungs intra-tracheally injected with AF-MSCs by culturing with a modification of small airway <em>growth</em> medium (mSAGM) lacking retinoic acid (RA) and triodothyronine (T3) with addition of <em>fibroblast</em> <em>growth</em> <em>factor</em>-10 (FGF10). Cells and scaffolds were characterized by immunofluorescence and RT-PCR for markers of viability, proliferation, and lung distal airway differentiation (TTF-1(+) and SPC(+)) in the absence of markers of brain (TuJ1(-)) and thyroid (Pax8(-)).

RESULTS

After culture in mSAGM on either Matrigel or lung scaffolds, there were TTF-1(+)/TuJ1(-)/Pax8(-) cells, indicating a lung precursor phenotype. In addition, SPC(+) cells also evolved suggesting a more mature lung phenotype.

CONCLUSIONS

We demonstrate that mid- to late-trimester AF-MSCs can be induced to develop into lung precursor cells when cultured on the appropriate extracellular matrix (ECM), making them a viable source for use in cell therapy or development of an ex vivo tissue engineered lung.

With increasing efforts to develop and utilize mouse models of a variety of neuro-developmental diseases, there is an urgent need for sensitive neuroimaging methods that enable in vivo analysis of subtle alterations in brain anatomy and function in mice. Previous studies have shown that the brains of <em>Fibroblast</em> <em>Growth</em> <em>Factor</em> <em>17</em> null mutants (Fgf<em>17</em>(-/-)) have anatomical abnormalities in the inferior colliculus (IC)-the auditory midbrain-and minor foliation defects in the cerebellum. In addition, changes in the expression domains of several cortical patterning genes were detected, without overt changes in forebrain morphology. Recently, it has also been reported that Fgf<em>17</em>(-/-) mutants have abnormal vocalization and social behaviors, phenotypes that could reflect molecular changes in the cortex and/or altered auditory processing / perception in these mice. We used manganese (Mn)-enhanced magnetic resonance imaging (MEMRI) to analyze the anatomical phenotype of Fgf<em>17</em>(-/-) mutants in more detail than achieved previously, detecting changes in IC, cerebellum, olfactory bulb, hypothalamus and frontal cortex. We also used MEMRI to characterize sound-evoked activity patterns, demonstrating a significant reduction of the active IC volume in Fgf<em>17</em>(-/-) mice. Furthermore, tone-specific (16- and 40-kHz) activity patterns in the IC of Fgf<em>17</em>(-/-) mice were observed to be largely overlapping, in contrast to the normal pattern, separated along the dorsal-ventral axis. These results demonstrate that Fgf<em>17</em> plays important roles in both the anatomical and functional development of the auditory midbrain, and show the utility of MEMRI for in vivo analyses of mutant mice with subtle brain defects.

The regional localization of five reference loci to sheep chromosomes is reported. The newly mapped loci are the T-cell receptor, beta (TCRB), coagulation <em>factor</em> X (F10), laminin gamma 1 (LAMC1), cyclic GMP rod phosphodiesterase, alpha (PDEA) and <em>fibroblast</em> <em>growth</em> <em>factor</em> 2 (FGF2). The assignments of PDEA and LAMC1 to chromosomes 5q23-q31 and 12q22-q24 respectively provide the first markers physically assigned to these chromosomes. They also allow the provisional assignment of sheep syntenic group U19 to chromosome 5 and U1 to chromosome 12. The mapping of FGF2 to chromosome <em>17</em>q23-q25 anchors the unassigned linkage group 'A' to chromosome <em>17</em>, and the assignment of TCRB to chromosome 4q32-qter facilitates the orientation of a linkage group on sheep chromosome 4. The mapping of F10 to sheep chromosome 10q23-qter supports the recent assignment of bovine syntenic group U27 to cattle chromosome 12, as sheep chromosome 10 and cattle chromosome 12 are banded homologues.