Hypochondroplasia (HCH) is caused by mutations in the <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor type 3 (FGFR 3). Prenatal diagnosis of HCH based exclusively on the sonographic measurements of the fetal skeleton is difficult and has not been reported. We describe a newborn infant with HCH who was born to a mother with achondroplasia (ACH) and a father with HCH. Serial sonographic measurements were recorded from 16 weeks of gestation. All measurements remained normal up to <em>22</em> weeks of gestation. At 25 weeks of gestation, the long bones began to appear shorter than expected for gestational age, while the head measurements (biparietal diameter and head circumference) remained normal. The measurements were sufficiently different to distinguish from findings in normal and achondroplastic fetuses. Our findings suggest that it is possible to distinguish the normal fetus from a fetus affected with HCH and to distinguish HCH and ACH from each other based on the sonographic measurements alone. To our knowledge, this is the first report of longitudinal sonographic measurements of HCH in the second and third trimesters.

OBJECTIVE

This study aimed to evaluate the associations between the concentrations of three major angiogenic cytokines-vascular endothelial growth factor-A165 (VEGF-A165), basic fibroblast growth factor (bFGF), and platelet-derived growth factor-BB (PDGF-BB)-in the tear of sulfur mustard (SM)-exposed subjects and corneal neovascularization (CNV) 26 years after exposure.

METHODS

The concentrations of VEGF-A, bFGF, and PDGF-BB were measured by enzyme-linked immunosorbent assay (ELISA) in reflex tears of (i) SM-injured patients with CNV (positive case group including 18 individuals) and (ii) SM-injured patients without CNV (negative case group including 22 individuals). Then results were compared to corresponding values obtained from tears of 40 healthy control subjects.

RESULTS

The mean concentrations of all investigated growth factors, VEGF-A165, bFGF, and PDGF-BB, were significantly higher in positive cases than controls (P ≤ 0.001, P = 0.028, and P = 0.041, respectively). Whereas, VEGF-A165 was the only growth factor which displayed significantly elevated concentrations in negative case group compared to the healthy individuals (P = 0.030). Additionally, the mean level of VEGF-A165 was also higher in positive patient group than negative patients (P = 0.022). Subjects with increased concentrations of tear VEGF-A165 were more than 10 times more likely to suffer from CNV than normal individuals (odds ratio (OR) = 10.43, confidence interval (CI): 2.14-38.46, P = 0.001), while elevated levels of bFGF and PDGF-BB increased the risk of CNV by about twofold.

CONCLUSIONS

Although all investigated cytokines had increased in tears of positive patients, VEGF-A was the only one which showed a significant correlation with the severity of CNV, and thus played a crucial role in corneal angiogenesis.

Basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) has shown a neurotrophic effect in the neurons of several CNS areas. In vivo, it contributes to restore neurochemical and morphological deficits in different rodent models of brain damage, including rats with brain damage induced by hypoxia/ischemia when FGF was intramuscularly (i.m.) administered. Toxicological and immunological studies performed in rats, mice and volunteers showed no evidence of side-effects. Bovine FGF was i.m. administered in children with mental retardation caused by perinatal hypoxia, aged 1-15 years, at dosages of 0.4 or 0.28 microgram kg-1, once or twice a month, over 7-12 months. Group A [n = 12; 6 treated (T), 6 controls (Ct)], group B (n = 16; 8 T, 8 Ct) and group C (n = 67; 45 T, <em>22</em> Ct) were evaluated with the P. A. R. scale, the WISC-RM and the Gesell scale, respectively. Development increased significantly in treated children from groups A (P < 0.02) and C (P < 0.001), and IQ rose by more than 10 points (P < 0.001) in group B patients.

Craniosynostosis is the premature fusion of the cranial vault sutures. We have previously described a colony of rabbits with a heritable pattern of nonsyndromic, coronal suture synostosis; however, the underlying genetic defect remains unknown. We now report a molecular analysis to determine if four genes implicated in human craniosynostosis, TWIST1 and <em>fibroblast</em> <em>growth</em> <em>factor</em> receptors 1-3 (FGFR1-3), could be the loci of the causative mutation in this unique rabbit model. Single nucleotide polymorphisms (SNPs) were identified within the Twist1, FGFR1, and FGFR2 genes, and the allelic patterns of these silent mutations were examined in <em>22</em> craniosynostotic rabbits. SNP analysis of the Twist1, FGFR1, and FGFR2 genes indicated that none were the locus of origin of the craniosynostotic phenotype. In addition, no structural mutations were identified by direct sequence analysis of Twist1 and FGFR3 cDNAs. These data indicate that the causative locus for heritable craniosynostosis in this rabbit model is not within the Twist1, FGFR1, and FGFR2 genes. Although a locus in intronic or flanking sequences of FGFR3 remains possible, no direct structural mutation was identified for FGFR3.

Augmentation of thrombin-modulated chemotaxis and mitogenic activity within the early phase of soft tissue repair is now possible. Identification of high-affinity thrombin receptor binding domains within thrombin has enabled the synthesis of a family of peptides which interact with thrombin receptors and enhance in vitro mitogenesis. A single (5.0 micrograms/wound) application of the thrombin receptor-activating peptide (P517-30) significantly increased wound breaking strength from Day 5 (31% over controls) to Day 12. Two models of impaired healing created by radiotherapy (RT) were used to elucidate possible mechanisms of P517-30 action. Although P517-30 did not completely overcome the RT-induced healing impairments, it increased breaking strength under conditions of penetrating whole body RT-induced pancytopenia by <em>22</em>% and of nonpenetrating surface RT-induced dermal cell damage by 42%. This suggests that P517-30 directly stimulates resident endothelial cells, <em>fibroblasts</em>, or other cells to overcome dermal and circulating monocytic deficits. These results suggest a method to accelerate wound healing with potential clinical applications and emphasize the activity of thrombin as a <em>growth</em> <em>factor</em>.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> 23 (FGF23) is a potent regulator of Pi and 1,25-(OH)(2)D homeostasis. Early postpartum infants show intriguing changes in serum levels of Ca, Pi, PTH and 1,25-(OH)(2)D. However, the role of FGF23 in the early neonatal mineral metabolism has not been clarified. In order to evaluate the significance of FGF23 during the early postpartum period, we examined the circulating FGF23 levels using an intact FGF23 ELISA and a C-terminal FGF23 ELISA either in <em>22</em> umbilical cord blood samples (the cord blood) or in <em>22</em> term infants at 5 days of life (the 5-day-old infant). We also compared these ranges with those of 11 healthy adults. Data were expressed as mean+/-SD, and analyzed by two-way ANOVA, followed by the Tukey's test. C-terminal FGF23 in the cord blood, the 5-day-old infants and the healthy adults were 73.3+/-<em>22</em>.4, 81.0+/-28.2 and 39.0+/-7.8 RU/ml, respectively. Intact FGF23 in the cord blood, the 5-day-old infants and the healthy adults were 3.9+/-1.6, 21.8+/-17.6, and 27.6+/-7.3 pg/ml, respectively. Immunoprecipitation assays using anti-FGF23 antibodies demonstrated that the intact 32 kDa FGF23 was low and the fragmented FGF23 of 18 kDa was abundant in the cord blood compared with those in the healthy adults. In conclusion, our observations indicated that the intact FGF23/C-terminal FGF23 ratio was very low due to the fragmentation of FGF23 during the early postpartum period and might have a considerable contribution to the Pi homeostasis in the healthy term infants.

Our previous studies showed that the amount of endogenous basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) was reduced after ischemia and reperfusion insult. One of the mechanisms involved in the decrease of endogenous bFGF is the increased destruction of this <em>growth</em> <em>factor</em> associated with oxygen free radical activation and inflammation. We hypothesized that the wounding also impairs the secretion of bFGF and examined the bFGF gene expression in skeletal muscles after ischemia and reperfusion insult. In this study, a rat leg ischemia (4 h) and reperfusion (24 h) injury model was prepared and the in situ hybridization method and reverse transcriptase polymerase chain reaction technique (RT-PCR) were used to evaluate the bFGF gene expression and its localization in control (normal) and injured rat skeletal muscles. The results showed that the bFGF mRNA expression was localized in the cytoplasm in control skeletal muscle, especially at the periphery inside the cells. According to the intensity of the stain, four main classes of fibers could be identified: strongly, moderately, weakly, and negatively stained fibers. Based on the positive stain, about 82% of the total fibers examined were positive for bFGF mRNA stain. In ischemic or ischemic and reperfused rat skeletal muscles, the localization of bFGF mRNA expression was similar to that in normal skeletal muscles, but only 52% in ischemic muscles and <em>22</em>% in ischemic and reperfused muscles had positive bFGF mRNA staining. RT-PCR confirmed a significant decrease in bFGF mRNA expression in ischemic and reperfused rat skeletal muscles. These results suggest that the acute ischemia and reperfusion not only induce the destruction of endogenous bFGF molecule, which is stored at the extracellular matrix of the fibers, but also downregulate the bFGF gene expression. The simultaneous dysregulation of endogenous bFGF gene expression and decreased synthesis of bFGF protein suggest a possible role of this <em>growth</em> <em>factor</em> in delayed wound healing.

Acidic <em>fibroblast</em> <em>growth</em> <em>factor</em> causes an acute and transient nitric oxide-dependent hypotensive effect in experimental animals. However, this response is not found, or is very small, in vitro. We hypothesized that plasma mediators, such as kinins, are involved in aFGF-induced hypotension. We studied the hypotensive effect of intravenous aFGF (1 microg) in control Wistar rats, and compared this response to that in Wistar rats treated with a bradykinin receptor antagonist Na-adamantaneacetyl-D-Arg-(Hyp3,Thi5,8,D-Phe7]-brad yki nin), in kininogen-deficient Brown-Norway-Katholiek (BNK) rats, and in rats depleted of kininogen after repeated treatment with ellagic acid. FGF was administered in the jugular vein and mean arterial pressure was measured through a femoral artery catheter. Following treatment with the bradykinin receptor antagonist, the hypotensive effect of aFGF was reduced 38% with 58 microg of antagonist and by 60% with the 420 microg dose (9 +/- 1 vs <em>22</em> +/- 3mm Hg, p<0.01). Mean blood pressure decrease was 12 +/- 1 in BNK rats (p<0.01, vs control) and 10 +/- 2 mm Hg in kininogen-depleted ellagic acid-treated rats (p<0.05, vs control). These findings implicate kinins as necessary mediators for the hypotensive effect of aFGF in vivo. A full hypotensive effect of aFGF requires sufficient amounts of kininogens, the precursor molecules of kinins, as well as bradykinin receptors.

Astrocytes react to all noxae which damage neurons. Their reactions include degeneration, hypertrophy, hyperplasia and fibre formation. <em>Growth</em> <em>factors</em> inducing proliferation and differentiation of both neurons and astrocytes in culture play a pivotal role in the dynamic flow of signaling molecules between neurons and astroglia. Estrogens as well influence astroglia and are neuroprotectants. This study has investigated the interactions between <em>growth</em> <em>factors</em> and estrogens on DNA labeling and cytoskeletal protein [glial fibrillary acidic protein (GFAP) and vimentin] expression in <em>22</em> DIV astrocyte cultures treated for 24 or 36 h under different experimental conditions. Contemporary addition of 17-beta-estradiol (E2) with two or three <em>growth</em> <em>factors</em> for 24 h, significantly stimulated methyl-[3H]thymidine incorporation into DNA from <em>22</em> days in vitro (DIV) astrocyte cultures. This effect reached a peak when E2 was co-added with epidermal <em>growth</em> <em>factor</em> (EGF), basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) and insulin. In astrocyte cultures treated for 36 h with E2 and EGF + insulin or bFGF + insulin added in the last 12 h, DNA labeling was remarkably increased. The parallel cyclin Dl expression positively correlated with ERK2 activation. Western blot analysis for cytoskeletal proteins showed also changes of both GFAP and vimentin expression. The above data suggest the occurrence of a scheduled interaction between "competence" or "progression" <em>growth</em> <em>factors</em> and estrogens on DNA labeling and cytoskeletal protein expression during astroglial cell proliferation and differentiation in culture. A better understanding of the mechanisms of these interactions may contribute to develop strategies for controlling astroglial reaction in cerebrovascular disease including stroke and hypertensive brain damage.

OBJECTIVE

To study the preventive effect of recombinant human basic fibroblast growth factor (rh-bFGF) on restenosis after percutaneous transluminal coronary angioplasty (PTCA).

METHODS

Sixty male Wistar rats were randomly divided into sham operation group, intima injured group, and intima injured plus rh-bFGF treated group. Rat carotid arteries were injured using a balloon catheter except sham operation group. The rats of rh-bFGF treated group were injected im rh-bFGF 10 kU . kg-1 . d-1 after intima was injured. Ten rats in each group were killed on d 7 and d 14 after injury, respectively. [3H]Thymidine incorporation assay and pathological examination were carried out to each vessel.

RESULTS

(1) Seven days after injury, the average intimal thickness in sham operation group, intima injured group, and rh-bFGF treated group was (7 +/- 1), (32 +/- 11), and (17 +/- 3) micron; average intimal area was (384 +/- 145), (1530 +/- 817), and (586 +/- 185) micron2; the numbers of smooth muscle cells in neointima per transect were 0 +/- 0, 146 +/- 18, and 105 +/- 26; the ratio of the collagen area to intimal area plus medial area were 0.29 +/- 0.09, 0.7 +/- 0.3, and 0.30 +/- 0.14; [3H]thymidine incorporation were (17 +/- 6), (62 +/- 23), (20 +/- 8) kBq/g tissue, respectively. (2) Fourteen days after injury, the average intimal thickness of sham operation group, intima injured group, and rh-bFGF treated group was (8 +/- 1), (41 +/- 9), and (20 +/- 3) micron; average intimal area was (391 +/- 134), (1761 +/- 337), and (731 +/- 124) micron2; the numbers of smooth muscle cells in neointima per transect were 0 +/- 0, 145 +/- 9, and 102 +/- 6; the ratio of collagen area to intimal area plus medial area were 0.28 +/- 0.14, 0.59 +/- 0.21, and 0.38 +/- 0.03; [3H]thymidine incorporation was (15 +/- 4), (57 +/- 11), and (22 +/- 6) kBq/g tissue, respectively.

CONCLUSIONS

Large dosage of rh-bFGF inhibits neointimal hyperplasia and reduces restenosis after balloon injury.

Head and Neck Squamous cell carcinoma (HNSCC) can be characterized by synchronous tumors in the upper aerodigestive tract. Second primary tumors as a result of field cancerization are a significant problem amongst patients with risk <em>factors</em> for HNSCC, indicating a need for chemo preventive agents. We investigated the efficacy of local and systemic Curcumin C3 complex (C3); a purified mixture of Curcumin, bisdemethoxy Curcumin and demethoxy Curcumin as a chemo preventative agent in 4-nitroquinoline-1-oxide (4NQO)-induced tumorigenesis in mice. The effect of local C3 application was compared to C3 administered orally and in combination with systemic administration. C57Bl/6 mice were administered 4NQO (50 µg/ml) in the drinking water for 16 weeks. At 12 weeks, mice were subjected to daily treatment with either vehicle (control), or 15 mg C3 complex by local delivery, gavage, or combined local and gavage for 28 days (16 week time point), and followed up to <em>22</em> weeks. Compared to local and oral systemic C3 administration, combination of local and systemic application significantly decreased multiplicity of 4NQO-induced preneoplastic and neoplastic lesions (p<0.05). Treatment with C3 correlated with a decrease in cell proliferation compared to the 4NQO group. Further, pre-treatment with C3 complex significantly attenuated 4NQO induced expression of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF-2) and its cognate receptor FGFR-2, suggesting an important role of FGF-2/FGFR-2 axis in chemoprevention of HNSCC (p<0.05). Our findings suggest that a combination of local and systemic C3 complex could effectively target proliferation and inhibit 4NQO-induced tumorigenesis <i>via</i> modulation of the FGF-2/FGFR-2 axis as a mechanism for its efficacy.

Gene expression patterns have been investigated in prolonged cultures of rat mandibular condylar cartilage (MCC) cells to examine the possibility of culture-induced phenotypic changes. MCC cells were isolated from newborn rats and grown in the presence of 10% fetal bovine serum (FBS) and basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF). MCC cells were passaged and cultured in the presence of 10% FBS and bFGF until confluent. After confluence, the medium was changed to that supplemented with 10% FBS, ascorbate, and beta-glycerophosphate (day 1). Mineralization and gene expression of MCC cells have been investigated. Mineralization, visualized by staining with Alizarin red, was observed to begin at day 13 in culture, and increased up to day <em>22</em> in culture, which was the length of this study. Type II collagen and aggrecan mRNAs were highly expressed at the start of culture (day 1-4) and decreased in a time-dependent manner. Type I collagen, alkaline phosphatase, and osteopontin mRNAs expressed biphasic patterns that peaked at the start of culture and the beginning of mineralization (day 13-16). Osteonectin mRNA was expressed throughout the culture period. Osteocalcin mRNA was expressed before the beginning of mineralization peaking at day 7. These observations suggest that the gene expression patterns of MCC cells can be categorized into two different periods in prolonged culture: maturation (day 1-10) and mineralization (day 13-<em>22</em>). The day culture system of MCC represents a new model system in which the differentiation of embryonic MCC cells can be examined.

In a previous work we have shown that culturing adult rat dorsal root ganglia neurons modifies their neurotransmitter phenotype in such a way that cultured neurons synthesize transmitters that are not found in situ, while several other transmitters are expressed in a much higher percentage of neurons in culture than in situ [Schoenen J. et al. (1989) J. Neurosci. Res. <em>22</em>, 473-487]. The aim of the present study was to investigate the origin and the nature of the relevant environmental signals that allow this plasticity to be expressed, focusing on three neurotransmitters: 5-hydroxytryptamine, thyrotropin-releasing hormone and calcitonin-gene related peptide. The main results can be summarized as follows: (1) culturing cells in fetal calf serum or on feeder layers of astrocytes, Schwann cells or <em>fibroblasts</em> partially inhibits the serotoninergic phenotype of dorsal root ganglia neurons; (2) in vivo disconnection of dorsal root ganglia from their spinal targets but not from their peripheral or supraspinal targets induces a significant increase of the percentage of 5-hydroxytryptamine- and thyrotropin-releasing hormone-positive neurons in disconnected ganglia; (3) <em>growth</em> <em>factors</em> such as ciliary neuronotrophic <em>factor</em> or basic <em>fibroblast</em> <em>growth</em> <em>factor</em> but not nerve <em>growth</em> <em>factor</em> repress 5-hydroxytryptamine and calcitonin gene-related peptide immunoreactivity in cultured sensory neurons. In conclusion, neurotransmitter gene expression of adult dorsal root ganglia neurons is controlled by complex influences. Our data suggest that thyrotropin-releasing hormone and 5-hydroxytryptamine gene expression are tonically repressed in vivo by <em>factors</em> originating from the spinal segmental level and that <em>growth</em> <em>factors</em> such as ciliary neurotrophic <em>factor</em> or basic <em>fibroblast</em> <em>growth</em> <em>factor</em> could be potential vectors of this repressing effect.

Transforming <em>growth</em> <em>factor</em>-alpha (TGF alpha) is a low molecular weight peptide that has been implicated in an autocrine <em>growth</em>-stimulation of tumor cell lines in vitro. Although its mRNA has been detected in biopsies from human malignancies, the importance of an autocrine mechanism for the <em>growth</em> of human malignancies in vivo is unknown. The purpose of our study was to determine the effects of recombinant human TGF alpha (rhTGF alpha) on the in vitro <em>growth</em> of fresh human tumors using a capillary cloning system. <em>Growth</em> was detected in control capillaries in 17 of 47 specimens (36%) without rhTGF alpha. Further stimulation of 12 of these tumors with rhTGF alpha obtained a maximal effect at 0.1 to 1 microgram/ml. Stimulation by rhTGF alpha was concentration-dependent with some tumors showing a biphasic response. Eight of <em>22</em> specimens not showing colony formation in control capillaries were recruited to form colonies by rhTGF alpha (p less than 0.005). Responsiveness to rhTGF alpha varied among individual tumors. In control experiments, rhTGF alpha failed to stimulate colony formation in three low-passage <em>fibroblast</em> cell lines. Furthermore, three additional human tumor specimens with benign cytologic findings displayed no colony formation. We concluded that a subgroup of primary human tumors was sensitive to rhTGF alpha in vitro.

Juvenile nasopharingeal angiofibroma (JNA) is a histologically benign locally aggressive tumor characterized by irregular vessels embedded in a fibrous stroma. Excessive vascularity results in bleeding complications, and the inhibition of angiogenesis is a promising strategy for managing extensive JNA tumors. To better characterize the endothelial components of JNA, we aimed to evaluate markers of vascular differentiation and proliferation, such as friend leukemia integration-1 (FLI-1) and endoglin, lymphatic markers, including podoplanin and vascular endothelial <em>growth</em> <em>factor</em> receptor 3 (VEGFR3) and its cognate ligand VEGFC, GLUT-1, a diagnostic marker that discriminates between hemangiomas and vascular malformations, and two markers of tissue remodeling, stromelysin 3 (ST3) and secreted acid protein rich in cysteine (SPARC). Antigens were assessed immunohistochemically in vessels and stromal cells of JNA archival cases (n=<em>22</em>). JNA endothelial cells were positive for endoglin, VEGFC and FLI-1, whereas podoplanin and VEGFR3 were negative in all cases. Both endothelial cells and <em>fibroblasts</em> stained for ST3 and SPARC. GLUT-1 was investigated in JNA cases, in infantile hemangiomas (n=123) and in vascular malformations (n=135) as controls. JNAs and vascular malformations were GLUT-1-negative, while hemangiomas showed positive staining. The presence of markers of endothelial differentiation and proliferation highlighted the hyper-proliferative state of JNA vessels. The absence of podoplanin and VEGFR3 underscores their blood endothelial cell characteristic. The absence of GLUT-1 discriminates JNAs from hemangiomas. ST3 and SPARC up-regulation in endothelial cells and <em>fibroblasts</em> may contribute to a compensatory signaling for controlling angiogenesis. Some of these markers may eventually serve as therapeutic targets. Our results may aid in the understanding of JNA pathophysiology.

Basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) is an angiogenic and mitogenic peptide that may have modulatory effects in the adult ovary and uterus. The FGF receptor is a tyrosine kinase and has similar affinity for both acidic FGF and bFGF. The ligand-binding portion of the receptor has three immunoglobulin-like domains. bFGF and the FGF receptor have been localized in the fetal rat ovary and Mullerian duct. bFGF is a proliferative agent recently shown to be vital for long term cultures of mouse fetal primordial germ cells. Expression of bFGF and FGF receptor mRNA have not heretofore been reported in human fetal ovary and uterus. We prepared RNA from whole human fetal ovaries and uteri at 10, 15, 19, and <em>22</em> weeks gestation. After reverse transcription of the RNA into cDNA, we used polymerase chain reaction (PCR) primers directed to specific portions of bFGF and the FGF receptor and performed PCR amplification using the fetal cDNA. We found mRNA expression of bFGF and two forms of FGF receptor in all fetal ovaries and uteri at these developmental stages. One of the PCR products for the FGF receptor was the sequence that contained three immunoglobulin-like domains, whereas a second PCR-amplified fragment was consistent with a FGF receptor mRNA that has a 267-basepair deletion in the first immunoglobulin-like loop. We conclude that bFGF and two forms of the FGF receptor mRNA are expressed in human fetal ovary and uterus. The finding that both bFGF and the FGF receptor are concurrently expressed suggests that bFGF may serve as an autocrine or paracrine modulator during early development of the human reproductive tract.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> 23 (FGF23)-induced hypophosphatemia is a rare paraneoplastic syndrome of phosphate wasting that, if unrecognized, may cause tumor-induced osteomalacia. It is classically associated with benign mesenchymal tumors but occasionally has been found in patients with other malignancies. Hypophosphatemia has been associated with acute leukemia but has not previously been reported to be due to inappropriate FGF23 secretion. Here, we describe FGF23-induced severe hypophosphatemia and renal phosphate wasting associated with a mixed-phenotype Philadelphia chromosome-like acute leukemia in a previously healthy <em>22</em>-year-old man. He was found to have low serum 1,25-dihydroxyvitamin D and extremely high FGF23 levels, as well as inappropriate urinary phosphorus excretion. The hypophosphatemia improved with calcitriol and oral phosphate treatment but normalized only during chemotherapy-induced ablation of the blasts. FGF23 levels declined with a reduction in peripheral blast counts. Using real-time reverse transcription polymerase chain reaction, we found that the leukemia cells were the source of FGF23. To our knowledge, this is the first description of FGF23-induced hypophosphatemia associated with acute leukemia. We recommend that the FGF23 paraneoplastic syndrome be considered as a possible etiology of hypophosphatemia in patients with acute leukemia.

Basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF2) is an important mediator of smooth muscle cell (SMC) proliferation following arterial injury that results in neointimal <em>growth</em>. The present study was designed to explore the effects of recombinant FGF2 linked to the ribosome-inactivating protein saporin-6 (rFGF2-SAP) on vascular SMC cytotoxicity and neointimal formation following arterial injury. Cultured rat aortic SMCs were exposed to various concentrations of rFGF2-SAP, FGF2, and saporin-6 (SAP). Incubation with rFGF2-SAP resulted in a decreased number of SMCs beginning at a concentration of 10(-9) mol/L. Significant cytotoxicity was observed with as little as a 30-minute exposure of SMCs to rFGF2-SAP. To evaluate the ability of rFGF2-SAP in an in vivo model to reduce neointimal formation, Sprague-Dawley rats underwent carotid artery balloon denudation and received an intravenous bolus of vehicle or 5, 10, 15, or 20 micrograms/kg rFGF2-SAP on 0, 3, 6, and 9 days after injury. Rats were euthanized at 14 days, and carotid arteries were analyzed by computerized morphometry. The threshold dose for a significant reduction in neointimal area by rFGF2-SAP was 15 micrograms/kg (47% reduction in neointima). When dosing was extended to include days 16, 19, and <em>22</em>, the neointima was reduced 33% at 28 days (P = .048). rFGF2-SAP reduced neointima without associated medial thinning or arterial wall dilatation. To determine if rFGF2-SAP directly targets SMCs in vivo, rats underwent carotid injury and received either 15 micrograms/kg rFGF2-SAP or vehicle on day 0 and at 72 hours, with euthanasia at 78 hours after balloon denudation. Medial SMC number was reduced 46% in the rFGF2-SAP group. Tissue sections from arteries 3 days after balloon injury demonstrated rFGF2-SAP binding to medial SMCs and adventitial cells. Staining for <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor 1 revealed a high level of expression in ballooned arteries 3 and 14 days after injury. Taken together, these results provide a molecular and cellular basis for the observed specificity. Prolonged delivery of rFGF2-SAP can affect the natural history of arterial repair after injury.

BackgroundAlthough aberrant proliferation and activation of lung <em>fibroblasts</em> are implicated in the initiation and progression of idiopathic pulmonary fibrosis (IPF), the underlying mechanisms are not well characterized. Numerous microRNAs (miRNAs) have been implicated in this process; however, miRNAs derived from exosomes and their relevance to <em>fibroblast</em>-to-myo<em>fibroblast</em> differentiation have not been fully elucidated. In this study, we aimed to identify exosome-derived miRNAs relevant to fibrosis development.MethodsWe profiled exosome-derived miRNAs expression in sera of C57BL/6 mice exhibiting bleomycin-induced pulmonary fibrosis by miRNA array analysis. After validating a selected miRNA by quantitative reverse-transcription polymerase chain reaction, its effect on <em>fibroblast</em>- to-myo<em>fibroblast</em> differentiation was investigated using human lung <em>fibroblasts</em>. Furthermore, we determined the role of the selected miRNA in an in-vivo pulmonary fibrosis model.ResultsMiRNA array analysis revealed that miR-<em>22</em> expression was increased by up to 2 fold on day 7 after bleomycin treatment compared with that in vehicle-treated mice. In vitro, miR-<em>22</em> transfectionsuppressed TGF-β1-induced α-SMA expression. This was mediated via the inhibition of the ERK1/2 pathway. Baseline α-SMA expression was increased upon miR-<em>22</em> inhibitor transfection. Furthermore, miR-<em>22</em> negatively regulated connective tissue <em>growth</em> <em>factor</em> expression in the presence of TGF-β1. In vivo, administration of a miR-<em>22</em> mimic on day 10 after bleomycin challenge ameliorated pulmonary fibrosis lesions accompanied by decreased α-SMA expression in the model mice.ConclusionsExosomal miR-<em>22</em> modulates <em>fibroblast</em>-to-myo<em>fibroblast</em> differentiation. The present study warrants further investigations to shed light on miR-<em>22</em> as a novel therapeutic target for patients with IPF.

OBJECTIVE

We examined serum concentrations of basic fibroblast growth factor (bFGF) in women with hypertensive disorders in pregnancy and analyzed whether serum concentrations of bFGF can be used as a discriminator between mild and severe preeclampsia.

METHODS

One hundred and twenty pregnant women were included in this prospective cohort study. We evaluated serum concentrations of bFGF in pregnant women with chronic hypertension (n=22), mild preeclampsia (n=40), severe preeclampsia (n=31), and healthy pregnant women (n=27).

RESULTS

Median serum concentrations of bFGF in healthy pregnant women, women with chronic hypertension, and women with mild or severe preeclampsia were 0.0 (0-37.2), 0.0 (0-3.0), 1.7 (0-97.2), and 0.0 (0-52.0), respectively. Comparison of the median values of serum bFGF concentrations showed a significant difference between healthy pregnant women and women with mild preeclampsia (P=0.02). In a logistic regression model, we found a significant influence of bFGF serum concentrations on the diagnosis of mild preeclampsia (P=0.01), but not on the diagnosis of chronic hypertension (P=0.19) or severe preeclampsia (P=0.41).

CONCLUSIONS

Elevated serum concentrations of bFGF are associated with mild preeclampsia, but are not discriminatory for the distinction between mild and severe preeclampsia.

BACKGROUND

The effectiveness of platelet-rich plasma (PRP) for treating soft tissue injuries is still controversial. Most of PRPs were prepared simply by concentrating in volume and were injected right after preparation in physician offices. Neither platelet count nor growth factors were quantitated in advance. We prepared and stored leukocyte and platelet-rich plasma (L-PRP) by regular separation protocols for blood components in the blood bank. And we investigated the dynamic change of growth factors in the L-PRPs over the period of storage.

METHODS

The L-PRPs were prepared by 2-step centrifugation and stored agitatedly at 22 °C for 7 days in the platelet incubator of blood bank. Levels of vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF)-basic, hepatocyte growth factor (HGF), insulin-like growth factor (IGF)-1, platelet derived growth factor (PDGF)-AB, endothelial growth factor (EGF), and transforming growth factor (TGF) over the period of storage were evaluated daily after freeze-thawing to release growth factors from platelet.

RESULTS

Compared to original whole blood, platelet concentration, VEGF, FGF-basic, PDGF-AB, EGF, and TGF-beta1 levels of L-PRPs significantly increased after PRP preparation. Both HGF and IGF-1 in L-PRPs remained the original plasma level. Platelet, FGF, and TGF-beta1 concentrations sustained during storage, and concentrations of VEGF, HGF, IGF-1, PDGF-AB, and EGF in L-PRPs increased over the period of storage.

CONCLUSIONS

During the storage in blood bank, platelet counts and 7 growth factors sustained or reached higher level than L-PRP obtained on first day. Multiple injections of stored PRPs could become applicable by our protocol.

The pharmacological manipulation of wound healing with locally applied <em>growth</em> <em>factors</em> is now a practical possibility. The effect of topical applications of recombinant basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) on the strength and cellularity of healing incisional rat skin wounds was investigated. Applications of bFGF in a simple vector (either a collagen suspension or saline) were not associated with any positive effects on wound breaking load at 7 days after injury in comparison with vector-treated control wounds; at the highest dose of 50 micrograms per wound, breaking loads were significantly decreased from a mean(s.e.m.) of 287(<em>22</em>) g/cm2 in controls to 201(23) g/cm2 (P < 0.005). Increasing doses of applied peptide were paralleled by increasing wound cellularity. Delay of bFGF release at the site of application was achieved by encapsulation into red blood cell ghosts. Wounds treated with bFGF in such ghosts were 50 per cent stronger than paired control wounds (388(27) versus 256(28) g/cm2, P < 0.002) 7 days after injury. Treated wounds were significantly more cellular at 4 days than paired control wounds. Topical applications of bFGF applied at the time of injury exert a positive effect on incisional wound strength only when a vector that delays release is used.

A method for culturing endothelial cells (HCC-EC) from surgical specimens of human corpus cavernosum has been developed. The approach involves selective endothelial out<em>growth</em> from explants and may be generally applicable to tissues whose endothelium is not amenable to isolation by routine mechanical or enzymatic methods. The tissue is minced into pieces which are placed onto gelatin- or fibronectin-coated tissue culture plastic, and grown in medium suitable for microvascular endothelial cell <em>growth</em> (Carson and Haudenschild, In Vitro <em>22</em>:344-354, 1986). By Days 5 to 7 EC colonies are found. Within a day or two after the appearance of the EC colonies, a non-EC cell type appears and, if undisturbed, quickly overgrows the EC. An exploitable temporal separation between the emergence of EC and non-EC is obtained when both conditioned medium (from bovine aortic endothelium) and retinal extract are present during the out<em>growth</em> period. Explants are removed by pipetting at the first sign of the emergence of the non-EC cell type. Once isolated, HCC-EC do not require conditioned medium but do require either retinal extract or acidic <em>fibroblast</em> <em>growth</em> <em>factor</em> for survival and <em>growth</em>. Approximately 60% of the first passage cultures are at least 80% EC as judged by DiI-Ac-LDL labeling. One corpus (0.3 x 0.3 x 0.5 cm) usually produces 120 cm2 of primary culture within 2 wk. These EC form contact-inhibited monolayers and stain positively for <em>Factor</em> VIII. They have a doubling time at 6th passage of 48 h and a plateau density of 5 to 7 x 10(4) cells/cm2. The availability of such cultures should facilitate the study of endothelium-mediated responses which play an important role in the erectile function of human penile corpus cavernosum.

Heparan sulfate proteoglycans are obligatory for receptor binding and mitogenic activity of the basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF). In the present study the influence of undersulfated heparan sulfate on the expression of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> and coronary smooth muscle cell (cSMC) proliferation was investigated. Chlorate, known to be an inhibitor of ATP-sulfurylase, was used as a tool to suppress sulfation of heparan sulfate. When cultured cSMC were treated with 10 mM sodium chlorate in sulfate-depleted medium, the cell number and [3H]thymidine incorporation decreased by 76% and 66% respectively, while the protein content per cell was doubled. At the same time the [35S]sulfate incorporation into cell-associated proteoglycans was reduced by 90%. The remaining minimal amount of available [35S]radioactivity was preferably incorporated into heparan sulfate. Under the same conditions the [6-(3)H]glucosamine incorporation into glycosaminoglycans was not impaired. The chlorate-induced increase of cell protein content includes an overexpression of bFGF, which increased from 6-8 ng to 18-<em>22</em> ng/mg cell protein. However, no changes in the distribution of bFGF between the intracellular and pericellular compartment could be observed. Cell cycle analysis by FACS revealed a G1 arrest of the cell cycle with increase of the G1/S ratio from 2.9 (control) to 6.1 (chlorate) but the DNA content per cell corresponded to normal diploid cells both in control and chlorate-treated cells. The chlorate effect can be abolished by addition of 5 mM sodium sulfate to the cultures. Our results demonstrate an inverse association between the sulfation of heparan sulfate and the expression of bFGF. They suggest that chlorate blocks the cell cycle in the late G1-phage and that mitogenesis of cSMC requires fully sulfated cell-associated proteoheparan sulfate.