The regulation of endothelin-B receptor (ETB) mRNA expression in human endothelial cells (ECs) by cytokines and <em>growth</em> <em>factors</em> may play an important role in the response of the endothelium to inflammatory and angiogenic stimuli. Using quantitative RT-PCR, we studied ETB expression in human umbilical vein ECs (HUVECs) grown in culture on either plastic or fibrin matrix for 24 h in the presence or absence of tumor necrosis <em>factor</em>-alpha (TNF-alpha, 100 U/ml) or basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF, 30 ng/ml). In addition, the effect of the nitric oxide (NO) donor S-nitrosyl-acetylpenicillamine (SNAP, 0.4 mM) was examined directly on ETB expression or on the response to bFGF. Under control conditions, ETB mRNA was detected after 35 cycles of amplification as a band of the expected size (553 bp). In the absence of fibrin matrix, ETB was downregulated by bFGF and TNF-alpha and could barely be detected by PCR. Southern analysis of the RT-PCR products after 25 cycles revealed that bFGF reduced ETB mRNA expression by 2.7 +/- 0.4-fold (p < 0.01) and TNF-alpha tended to reduce its expression by 1.8 +/- 0.9-fold of control, although this did not reach statistical significance (p < 0.20). In contrast, on fibrin matrix both bFGF and TNF-alpha increased ETB mRNA expression by 25 +/- 9-fold (p < 0.05) and 68 +/- <em>19</em>-fold (p < 0.05) of control, respectively, suggesting a role for ETB in the vascular tube formation that occurs under these conditions. Pharmacologic addition of NO mimicked the effect of fibrin, converting the response to bFGF from down- to upregulation of ETB, raising the possibility that NO acts as a molecular switch modulating the response to angiogenic <em>factors</em>.

OBJECTIVE

Apart from the constitutively activating mutation of the G-protein alpha subunit (Gsalpha) (gsp mutation), factors involved in tumorigenesis or those in tumour behaviour remain elusive in sporadic GH-secreting pituitary adenomas. Recently, the N-terminally truncated form of fibroblast growth factor receptor-4 (ptd-FGFR4) was identified in pituitary adenomas. This aberrant receptor has transforming activity, and causes pituitary adenomas in transgenic mice. The clinical relevance of this receptor warrants investigation. Our objective was twofold: first, to examine how the expression of ptd-FGFR4 relates to gsp mutations; and second, to see whether patients with this receptor have unique clinical characteristics.

METHODS

mRNA was extracted from excised adenomas of 45 Japanese acromegalic patients. ptd-FGFR4 expression and gsp mutations were determined by reverse transcription polymerase chain reaction (RT-PCR) and direct sequencing. Preoperative clinical data were collected by reviewing medical charts and pituitary magnetic resonance imaging (MRI) scans.

RESULTS

ptd-FGFR4 mRNA expression was detected in 19 out of 45 tumours (42.2%) while gsp mutations were detected in 25 out of 45 tumours (55.6%). The prevalence of ptd-FGFR4 expression did not differ between gsp-positive (44.0%) and gsp-negative (40.0%) tumours (P = 1.00). ptd-FGFR4-positive tumours invaded the cavernous sinus more frequently (P = 0.0098) than did the ptd-FGFR4-negative tumours. Tumour size was not statistically different between ptd-FGFR4-positive and -negative tumours (P = 0.198). The presence of ptd-FGFR4 did not correlate with age at operation, sex, preoperative serum GH or IGF-1 levels.

CONCLUSIONS

We found that ptd-FGFR4 expression and gsp mutations occur independently of each other, and that ptd-FGFR4 expression is associated with more invasive tumours in patients with GH-secreting pituitary adenomas.

By using a highly specific and sensitive heterologous radioimmunoassay, we determined the content of basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) in fresh samples of aqueous humor obtained from human (n = 18), feline (n = 8), canine (n = 12), and porcine (n = 10) eyes by paracentesis. The content of bFGF in human aqueous humor ranged from 0.480 to 1.44 ng ml-1 (mean +/- S.D. = 1.074 +/- 0.297 ng ml-1); in feline samples, from 0.672 to 1.152 ng ml-1 (1.016 +/- 0.158 ng ml-1); in canine samples, from 0.640 to 1.232 ng ml-1 (1.026 +/- 0.171 ng ml-1); and in porcine samples, from 0.627 to 0.957 ng ml-1 (0.728 +/- 0.115 ng ml-1). These values were confirmed by means of a dot/slot-blot technique. For all species, the aqueous samples had normal protein levels that ranged from 5 to <em>19</em> mg dl-1. There was no correlation of the content of bFGF with the level of protein or with age of the human subjects. The similarity in the concentrations of bFGF in the aqueous humor as well as the stability of the blood-aqueous barriers of all four species indicate that cats, dogs, and pigs can serve as suitable animal models for the study of the role of bFGF in health and disease. We suggest the possible involvement of bFGF in the pathogenesis of anterior-segment disorders, such as neovascular glaucoma, and in the wound-healing response of limbal tissues after glaucoma filtration surgery.

Recent reports indicate that glycosphingolipids play an important role in regulation of carbohydrate metabolism. We have shown that the iminosugar N-(5'-adamantane-1'-yl-methoxy)-pentyl-1-deoxynojirimycin (AMP-DNM), an inhibitor of the enzyme glucosylceramide synthase, is a potent enhancer of insulin signaling in rodent models for insulin resistance and type 2 diabetes. In this study, we determined whether AMP-DNM also affects lipid homeostasis and, in particular, the reverse cholesterol transport pathway. Treatment of C57BL/6J mice with AMP-DNM for 5 weeks decreased plasma levels of triglycerides and cholesterol by 35%, whereas neutral sterol excretion increased twofold. Secretion of biliary lipid also increased twofold, which resulted in a similar rise in bile flow. This effect was not due to altered expression levels or kinetics of the various export pumps involved in bile formation. However, the bile salt pool size increased and the expression of Cyp7A1 was up-regulated. In vitro experiments using HepG2 hepatoma cell line revealed this to be due to inhibition of <em>fibroblast</em> <em>growth</em> <em>factor</em>-<em>19</em> (FGF<em>19</em>)-mediated suppression of Cyp7A1 via the FGF receptor.

CONCLUSIONS

Pharmacological modulation of glycosphingolipid metabolism showed surprising effects on lipid homeostasis in C57BL/6J mice. Upon administration of 100 mg AMP-DNM/kg body weight/day, plasma cholesterol and triglyceride levels decreased, biliary lipid secretion doubled and also the endpoint of reverse cholesterol transport, neutral sterol excretion, doubled.

The present investigation was undertaken in order to study the long-term effects of perinatal asphyxia on basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF) gene expression and the number of dopamine nerve cell bodies in the mesencephalon of the rat. Asphyxia was induced during birth for <em>19</em>-20 min. A 30% increase in the number of tyrosine-hydroxylase immunoreactive (TH-IR) nerve cell bodies (i.e. dopamine-containing neurones) as well as a 50% increase in bFGF gene expression following asphyxia was found in the substantia nigra/ventral tegmental area 4 weeks after birth. The increase in bFGF mRNA levels may underlie the increase found in the number of dopamine cell bodies. The present results indicate that asphyxia during birth can prime the long-term development of the central nervous system.

Ovarian thecal cells are thought to differentiate from <em>fibroblast</em>-like precursor cells in the stroma adjacent to developing follicles. Since the precursor cells do not contain LH receptors, a regulator other than LH must initiate thecal differentiation. These studies were designed to test the hypothesis that preantral follicles secrete substances that can stimulate thecal differentiation. Preantral follicles devoid of theca were obtained by limited enzymatic dispersal of 26-day old rat ovaries. Follicles were cultured (5 follicles/well) in 96-well plates containing serum-free medium to generate follicle-conditioned medium (FCM). Isolated theca-interstitial cells (TIC) were cultured (2 days) in 50% FCM, to bioassay for androgen-stimulating activity. Androsterone production was measured by RIA. FCM from follicles of increasing size with 1, 2, 3, 4 or 5 layers of granulosa cells (GC) stimulated increasing amounts of androsterone suggesting that secretion of androgen-stimulating activity is developmentally regulated in preantral follicles. The androgen-stimulating activity of 7.5-fold concentrated FCM was markedly increased above control levels or the levels stimulated by insulin-like <em>growth</em> <em>factor</em>-I (100 ng/ ml), transforming <em>growth</em> <em>factor</em>-α (100 ng/ml), transforming <em>growth</em>-<em>factor</em>-β1 (10 ng/ml), inhibin A (300 ng/ml), activin A (100 ng/ml), or Müllerian inhibiting substance (MIS; 300 ng/ml) suggesting that the bioactive substances were distinct from these intrafollicular <em>growth</em> <em>factors</em>. rFSH stimulated a>> 10-fold increase in androgenstimulating activity demonstrating that the bioactivity is hormonally regulated. The bioactivity was sensitive to trypsin digestion but was not inhibited by indomethacin (10 μm) suggesting that it is peptide not prostaglandin in nature. Gel filtration chromatography indicated that the M of the bioactive peptides in FCM ranged from <em>19</em> 500 to 23 600. Taken together our results demonstrate that preantral follicles secrete thecal differentiating <em>factors</em> (TDFs) that are developmentally and hormonally regulated by FSH. The properties of the TDFs are markedly different from known intrafollicular <em>growth</em> <em>factors</em> and may represent a new paracrine regulator in the ovary that can stimulate LH-independent thecal differentiation.

Imatinib mesylate is a specific tyrosine kinase inhibitor that may block the platelet-derived <em>growth</em> <em>factor</em> and transforming <em>growth</em> <em>factor</em> pathways. These pathways are known to provoke <em>fibroblast</em> activation. We evaluated whether imatinib, by inhibiting these pathways, prevents diastolic dysfunction and attenuates myocardial remodeling using spontaneously hypertensive rats (SHRs). Eight-week-old male SHRs were randomly assigned to either imatinib treatment group (30 mg/kg per day; n=10; SHR-I) or hypertensive control group (distilled water, n=10; SHR-C). Wistar-Kyoto rats were used as normal controls (n=10). At 16 weeks, all rats underwent hemodynamic studies and Doppler echocardiography and then were euthanized. Their hearts were extracted for histopathologic, immunoblotting, and quantitative reverse transcriptase polymerase chain reaction analyses. Although imatinib did not affect blood pressure, it markedly reduced perivascular and interstitial fibrosis in the hearts of SHR. Echocardiogram showed that imatinib significantly reduced the left ventricular wall thickness (septal/posterior wall; SHR-C versus SHR-I, 18±1/<em>19</em>±2 versus 15±1/15±1 mm; P<0.001) and increased the E/A ratio (SHR-C versus SHR-I, 1.59±0.11 versus 1.84±0.16; P=0.001). Also, imatinib significantly reduced the mRNA expression of collagen type I, III, and platelet-derived <em>growth</em> <em>factor</em> receptor-β phosphorylation in the hearts of SHR. In addition, imatinib reduced collagen production by inhibiting the phosphorylation of c-abl and platelet-derived <em>growth</em> <em>factor</em> receptor-β in rat cardiac <em>fibroblasts</em>. In conclusion, these results suggest that imatinib could attenuate myocardial remodeling and improve left ventricular diastolic dysfunction in a hypertensive rat model by affecting platelet-derived <em>growth</em> <em>factor</em> and transforming <em>growth</em> <em>factor</em>-β1 pathway without the blood pressure-lowering effect.

The intestinal endocrine hormone human <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>) is involved in the regulation of not only hepatic bile acid metabolism but also carbohydrate and lipid metabolism. In the present study, bile acid/farnesoid X receptor (FXR) responsiveness in the FGF<em>19</em> promoter region was investigated by a reporter assay using the human colon carcinoma cell line LS174T. The assay revealed the presence of bile acid/FXR-responsive elements in the 5'-flanking region up to 8.8 kb of FGF<em>19</em>. Deletion analysis indicated that regions from -1866 to -1833, from -1427 to -1353, and from -75 to +262 were involved in FXR responsiveness. Four, four, and two consecutive half-sites of nuclear receptors were observed in the three regions, respectively. An electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay revealed FXR/retinoid X receptor α (RXRα) heterodimer binding in these three regions. EMSA and reporter assays using mutated constructs indicated that the nuclear receptor IR1, ER2, and DR8 motifs in the 5'-flanking region were involved in FXR responsiveness of FGF<em>19</em>. Lithocholic acid (LCA) (10 μM), chenodeoxycholic acid (CDCA) (10 μM), or GW4064 (0.1 μM) treatment increased reporter activity in a construct including the three motifs under FXR-expressing conditions whereas LCA and not CDCA or GW4064 treatment increased the reporter activity under pregnane X receptor (PXR)-expressing conditions. These results suggest that FGF<em>19</em> is transcriptionally activated through multiple FXR-responsive elements in the promoter region.

Subcutaneous implantation of demineralized bone particles (DBP) into rats induces the formation of a bone ossicle by a tightly controlled sequence of chondro- and osteo-inductive events which are directly comparable to those which occur in normal endochondral bone development. Although the morphological and biochemical sequence associated with endochondral bone formation in this model has been well characterized, to date little information is available as to the gene regulation by which these events occur. To examine the expression of genes in this system, RNA was isolated from implants every 2 days over a time course spanning 3 to <em>19</em> days after implantation of DBP into rats. Cellular levels of mRNA transcripts of cell-<em>growth</em>-regulated and tissue-specific genes were examined by slot blot analysis and compared to the morphological changes occurring during formation of the ossicle. Analysis of the mRNA levels of histone H4 and c-myc, markers of proliferative activity, revealed several periods of actively proliferating cells, corresponding to 1) production of fibroprogenitor cells (day 3), 2) onset of bone formation (day 9), and 3) formation of bone marrow (day <em>19</em>). The mRNA levels of collagen type II, a phenotypic marker of cartilage, peaked between days 7 and 9 post-implantation, corresponding to the appearance of chondrocytes in the implant, and rapidly declined on day 11 (to 5% of maximum value) when bone formation was observed. The peak mRNA levels of collagen type I, found in <em>fibroblasts</em> and osteoblasts, occurred first with the onset of bone formation (days 7-10) and again during formation of bone marrow (day <em>19</em>). This study has demonstrated that the temporal patterns of mRNA expression of cartilage type II and bone type I collagens coincide with the morphological sequence in this model of endochondral bone formation. Further, the mRNA levels of transforming <em>growth</em> <em>factor</em> beta 1 (TGF beta) were compared to those of collagen types I and II; a direct temporal correlation of TGF beta mRNA levels with that of collagen type I was found throughout the developmental time course. This observation of a tightly coupled relationship between TGF beta and type I collagen mRNA levels is consistent with a functional role for TGF beta in extracellular matrix production during in vivo bone formation.

OBJECTIVE

Dovitinib is a small molecule kinase inhibitor with activity against the fibroblast growth factor and vascular endothelial growth factor receptor families. The purpose of this phase Ib study was to define the recommended phase 2 dose of the combinations of gemcitabine and cisplatin or gemcitabine and carboplatin plus dovitinib.

METHODS

Patients with advanced solid tumors were enrolled in two parallel dose escalation arms (cisplatin- or carboplatin-based regimens). Treatment was administered with gemcitabine (1,000 mg/m(2) on days 1 and 8), cisplatin (70 mg/m(2)), or carboplatin (AUC 5) on day 1, and dovitinib (orally on days 1-5, 8-12, and 15-19), every 21 days. The starting dose of dovitinib was 300 mg and was dose escalated in successive cohorts using 3 + 3 dose escalation rules.

RESULTS

Fourteen patients with advanced solid tumors were enrolled, five to the cisplatin arm and nine to the carboplatin arm. Patients enrolled in the cisplatin arm received a median of two cycles of treatment (range 1-5), and patients enrolled in the carboplatin arm received a median of one cycle of treatment (range 1-4). There were no protocol-defined dose-limiting toxicities in the cisplatin arm. However, the cohort was closed due to the need for frequent dose delays and/or reductions and two patients experiencing severe thromboembolic events. There were two dose-limiting toxicities in the carboplatin arm at the starting dose level of dovitinib (both prolonged neutropenia), and the dose of dovitinib was de-escalated to 200 mg. Two additional dose-limiting toxicities (prolonged neutropenia and febrile neutropenia) occurred in the lower dose cohort, and the study was closed. No patients achieved an objective response to treatment.

CONCLUSIONS

Dovitinib in combination with gemcitabine plus cisplatin or gemcitabine plus carboplatin was poorly tolerated due to myelosuppression.

The biological heterogeneity of breast cancer leads to the need for finding new approaches to understand the mechanisms implicated in breast cancer progression. The tumor stroma appears as a key in the progression of solid tumors towards a malignant phenotype. Cancer associated <em>fibroblasts</em> (CAFs) may orchestrate a functional "corrupted" stroma which in turn helps metastatic spread. In this study, we investigated by real-time PCR, the expression of <em>19</em> <em>factors</em> by normal breast-associated <em>fibroblasts</em> (NAFs) and CAFs, which were implicated in several actions promoting tumor <em>growth</em>, such as extracellular matrix remodeling, inflammation and invasion. Also, we explored the influence of inflammatory cells phenotypes (MMP11 status) and breast cancer cell lines (MCF-7 and MDA-MB-231) on the molecular profile of CAFs. If we consider that one of the major sources of CAFs are resident NAFs, the transition of NAFs into CAFs is associated with molecular changes involving the overexpression of some molecular <em>factors</em> of biological importance in tumor progression. In addition, the characterization of the tumor stroma regarding to the MMP11 status by MICs reflects a type of <em>fibroblasts</em> which contribute even more to tumor progression. Moreover, different patterns in the induction of the expression of <em>factors</em> by CAFs were observed, depending on the tumor cell line which they were co-cultured with. Furthermore, CAFs influence TGFβ expression in both cancer cell lines. Therefore, this study can help to a better characterization of tumor stroma in order to improve the prognostic evaluation, as well as to define the different populations of CAFs as potential therapeutic targets in breast cancer. © 2015 Wiley Periodicals, Inc.

OBJECTIVE

Bile acid synthesis is regulated by nuclear receptors including farnesoid X receptor (FXR) and small heterodimer partner (SHP), and by <em>fibroblast</em> <em>growth</em> <em>factor</em> 15/<em>19</em> (FGF15/<em>19</em>). We hypothesized that hepatic cysteine sulfinic acid decarboxylase (CSAD) (a key enzyme in taurine synthesis) is regulated by bile acids (BA). The aim of this study was to investigate CSAD regulation by BA dependent regulatory mechanisms.

METHODS

Mice were fed a control diet or a diet supplemented with either 0.5% cholate or 2% cholestyramine. To study BA dependent pathways, we utilized GW4064 (FXR agonist), FGF<em>19</em> or T-0901317 (liver X receptor [LXR] agonist) and Shp-/- mice. Tissue mRNA was determined by quantitative reverse transcription polymerase chain reaction. Amino acids were measured by high-performance liquid chromatography.

RESULTS

Mice supplemented with dietary cholate exhibited reduced hepatic CSAD mRNA while those receiving cholestyramine exhibited increased mRNA. Activation of FXR suppressed CSAD mRNA expression whereas CSAD expression was increased in Shp-/- mice. Hepatic hypotaurine concentration (the product of CSAD) was higher in Shp-/- mice with a corresponding increase in serum taurine conjugated BA. FGF<em>19</em> administration suppressed hepatic cholesterol 7-α-hydroxylase (CYP7A1) mRNA but did not change CSAD mRNA expression. LXR activation induced CYP7A1 mRNA yet failed to induce CSAD mRNA expression.

CONCLUSIONS

BA regulate CSAD mRNA expression in a feedback fashion via mechanisms involving SHP and FXR but not FGF15/<em>19</em> or LXR. These findings implicate BA as regulators of CSAD mRNA via mechanisms shared with CYP7A1.

Study Type - Therapy (case series) Level of Evidence 4 What's known on the subject? and What does the study add? Peyronie's disease (PD) is an acquired curvature of the penis attributable to progressive fibromatosis of the tunica albuginea (TA). It is frequently associated with Dupuytren's contracture and those of Ledderhose. More recently it was found that patients suffering from PD also often suffer from diabetes mellitus and gout. Cigarette smoking and the intake of large amounts of alcohol are considered risk factors for PD.The exact aetiology of the disease is unknown, however, the trauma hypothesis is shared by most authors. According to this theory, repeated sexual microtrauma in people genetically predisposed could cause PD. The inflammatory process leads to the formation of fibrosis and plaques. Plaque can lead to penile curvature and may reduce its functionality. Pain is the most common symptom of early-stage disease. In the late stages the pain disappears, but erectile dysfunction may occur. Surgical treatment is available, but this exposes the patient to a greater risk of erectile dysfunction and it is most frequently associated with a reduction in the length of the penis. The rationale for local medical therapy is to use a treatment that acts on the initial phase of the disease by reducing and stopping the processes that lead to fibrosis, thus stabilizing the disease. Systemic medical therapy is usually accompanied by high rates of recurrence. Many authors consider local drug therapy more appropriate. Local treatment consists of several types of medication, but results are often sub-optimal. Anti-inflammatory or immunoregulatory therapy, either systemic or topical, has shown some efficacy when administered early in the disease by modulating the inflammatory response and attenuating the alteration of tissue repair. Unfortunately, in most cases, patients are first seen when the plaque is chronically inflamed, stabilized and sometimes already calcified. We have tested a biological drug for intralesional administration for the first time. We chose iloprost, an analogue of prostacyclin I2, for its theoretically favourable properties. If used i.v., it has been shown to be effective in treating vascular ischaemic disease such as thromboangioitis obliterans, peripheral arterial occlusive disease, Raynaud's phenomenon and systemic sclerosis. The rationale for the therapeutic use of iloprost in the late stages of PD is based on the assumption that activation of fibrinolysis induced by the drug would be able to determine a regression of the plaque with a consequent reduction of the curvature on erection. The main purpose of this phase I study was to evaluate the safety and tolerability of this drug injected in the context of the fibrous plaque on a small number of patients before designing a large-scale randomized trial. According to the results,therapy with intralesional iloprost in Peyronie's disease seems to be safe and tolerated and is a possible alternative to surgery.

OBJECTIVE

To assess the safety and tolerability of intralesional injections of iloprost (an I2 prostacyclin analogue) for its ability to suppress the production of connective tissue growth factor in fibroblasts, for the treatment of Peyronie's disease (PD).

METHODS

A total of 38 patients with PD were preliminarily assessed according to symptoms, the degree of penile curvature and the size and number of plaques. Each patient received weekly intralesional injections of iloprost 200 ng in 1 mL normal saline for 5 weeks. If tolerated, the single dose was increased weekly to the maximum of 400 ng (2 mL). All the patients were preliminarily evaluated using at-home photography and were re-evaluated 2 months after the end of the treatment regimen. There was no placebo control group in this phase I study.

RESULTS

Almost all patients showed mild side effects (burning or pain) during the treatment at the site of injection. All patients tolerated well a iloprost dose of 200 ng; 19 patients reached a 300 ng dose and 14 tolerated a 400 ng dose without showing side effects. A total of 29% of the patients showed an improvement in curvature.

CONCLUSIONS

The results show that therapy with intralesional iloprost is a possible alternative to surgery for Peyronie's disease.

OBJECTIVE

This study aimed at investigating the relationship between the occurrence of uterine dehiscence in term pregnant scarred uteri and the presence of altered biochemical behavior of the scarring process.

METHODS

Collagen content and the expression of transforming <em>growth</em> <em>factor</em>-beta and its isoforms transforming <em>growth</em> <em>factor</em>-beta1 and transforming <em>growth</em> <em>factor</em>-beta3, connective tissue <em>growth</em> <em>factor</em>, basic <em>fibroblast</em> <em>growth</em> <em>factor</em>, vascular endothelial <em>growth</em> <em>factor</em>, platelet-derived <em>growth</em> <em>factor</em>, and tumor necrosis <em>factor</em>-alpha in myometrium of lower uterine segment were assessed in <em>19</em> otherwise healthy term patients with one previous cesarean delivery who were not in labor. We were searching for differences between patients who showed uterine dehiscence (9 cases) and patients who showed a normal-appearing scarred lower uterine segment (10 cases). We also evaluated all these features in lower uterine segment from unscarred uteri of 10 otherwise healthy patients who were not in labor.

RESULTS

In the case of uterine dehiscence, the scarred lower uterine segment showed a higher collagen content, a reduction of pan transforming growth factor-beta expression because of a marked decrease or absence of transforming growth factor-beta3, a reduction of connective tissue growth factor, an increase in basic fibroblast growth factor and a slight enhancement in vascular endothelial growth factor, platelet-derived growth factor, and tumor necrosis factor-alpha expression.

CONCLUSIONS

These findings contribute to meliorate our knowledge about uterine scar healing and allow us to hypothesize that uterine dehiscence of a scarred uterus may be related to altered biochemical behavior of the scarring process.

Vascular tumours are common lesions of the skin and subcutaneous tissue, but also occur in many other tissues and internal organs. The well-differentiated tumours consist of irregular anastomosing, blood-filled vascular channels that are lined by variably atypical endothelial cells. The less differentiated tumours may show solid strands and sheets, resembling carcinoma or lymphoma. Several <em>growth</em> <em>factors</em>, including basic <em>fibroblast</em> <em>growth</em> <em>factor</em>, transforming <em>growth</em> <em>factors</em> and vascular endothelial <em>growth</em> <em>factor</em>, play a role in tumour angiogenesis. <em>Growth</em> hormone (GH) is mitogenic for a variety of vascular tissue cells, including smooth muscle cells, <em>fibroblasts</em> and endothelial cells and exerts its regulatory functions in controlling metabolism, balanced <em>growth</em> and differentiated cell expression by acting on specific membrane-bound receptors, which trigger a phosphorylation cascade resulting in the modulation of numerous signalling pathways and of gene expression. Essential to the initiation of a cellular response to GH, the presence of receptors for this hormone may predict the adaptation of tumour cells resulting from GH exposure. To address the site/mode of action through which GH exerts its effects, a well characterized monoclonal antibody, obtained by hybridoma technology from Balb/c mice immunized with purified rabbit and rat liver GH-receptor (GHR) and directed against the hormone binding site of the receptor, was applied, using the ABC technique to determine GHR expression in a panel of vascular tumours. The GHR was cloned from a rabbit liver cDNA library with the aid of an oligonucleotide probe based on a <em>19</em> residue tryptic peptide sequence derived from 5900 fold purified rabbit liver receptor. A total of 64 benign and malignant vascular tumours were obtained from different human organ sites, including the chest wall, skin, axillary contents, duodenum, female breast, abdomen, stomach, colon, lymph node, bladder, body flank and neck regions. The tumours were of the following pathological entities: Haemangioma (n = 12); haemangioendothelioma (n = 10); Castleman's disease (n = 3), haemangiopericytoma (n = 4); angiosarcoma, (n = 11), Kaposi's sarcoma with focal infiltration by lymphoma, HIV +ve (n = 7), Kaposi's sarcoma (n = 17). The endothelial cell marker CD-31 was used to establish endothelial cell characteristics and microvascular density. To delineate tumour cell <em>growth</em>, immunohistochemical analysis of cycling nuclear protein and of proliferating cell nuclear antigen, using Ki-67 and PCNA polyclonal antibodies respectively, was used to demonstrate proliferative indexes. Results show that, compared to their normal tissue counterparts, nuclear and cytoplasmic expression of GHR consistently result in strong receptor immunoreactivity in the highly malignant angiosarcomas and Kaposi's sarcomas and was localized in the cell membranes and cytoplasm, but strong nuclear immunoreactivity was also identified. The presence of intracellular GHR is the result of endoplasmic reticulum and Golgi localization. Nuclear localization is due to identical nuclear GHR-binding protein. Furthermore, there was a positive correlation of GHR immunoreactivity with neoplastic cellular proliferation and cycling, as measured by Ki-67 and PCNA. In conclusion, this study shows that GHR expression in vascular tumours is a function of malignancy and cancer progression. Malignant cells, which are highly expressive of the receptor, have a greater proliferation rate and thereby also higher survival rate compared to tumours expressing lower or minimal receptor level. The presence of GHR in endothelial cells of vascular neoplasm indicates that they are target cells and GH is of importance in the proliferation of vascular tumour angiogenesis. GH is necessary not only for differentiation of progenitor cells, but also for their subsequent clonal expansion and maintenance. The results support the hypothesis that GH is involved in the paracrine-autocrine mechanism, acting locally in regulating vascular tumour <em>growth</em> and will be useful for site-specific studies of the evolution of vascular cancers. The use of anti-GHR antibodies to block tumour progression is an intriguing possibility.

This study assessed the grade of bone marrow (BM) fibrosis and its association with a seromarker for collagen-III formation and fibrosis-related cytokines in 25 immune thrombocytopenia (ITP) patients treated with thrombopoietin receptor agonists (Tpo-RA) who had at least one BM biopsy. Assessment of 8 pre- and on-treatment BM biopsies revealed statistically significant increases in reticulin. Reticulin in biopsies performed after a median of 1·4 years of treatment was graded: MF-0 in 3 (12%), MF-1 in <em>19</em> (76%), MF-2 in 2 (8%) and MF-3 in 1 (4%). No cytogenetic or flow-cytometric abnormalities were detected. Median pretreatment Procollagen III N-propeptide (PIIINP) (6·6 μg/l) was significantly higher than on-treatment levels (5·6 μg/l); both were higher than controls (3·4 μg/l; P < 0·001). PIIINP was negatively correlated with treatment duration (r = -0·49) suggesting a decelerated reticulin production over time. There was a trend towards an association between grade of reticulin and PIIINP. Transforming <em>growth</em> <em>factor</em> (GF)-beta and basic-<em>Fibroblast</em> GF were not different between patients and controls but Hepatocyte GF (HGF), an anti-fibrotic cytokine, was significantly elevated in patients. In conclusion, low-grade BM reticulin fibrosis is seen in most ITP patients on Tpo-RA. The novel findings of decreasing PIIINP and elevated HGF need further investigation to explore their significance in BM fibrogenesis.

BACKGROUND

TSU-68 is a novel multiple tyrosine kinase inhibitor that inhibits vascular endothelial growth factor receptor-2, platelet-derived growth factor receptor, and fibroblast growth factor receptor. This open-label, non-comparative, multicenter phase II study evaluated TSU-68 in combination with docetaxel in patients with metastatic breast cancer that had relapsed within 1 year despite prior treatment with an anthracycline-containing regimen.

METHODS

TSU-68 was orally administered on days 1-21, and docetaxel was intravenously delivered on day 1. The regimen was repeated every 21 days. Primary endpoint was objective response rate according to the RECIST guidelines version 1.0.

RESULTS

TSU-68 in combination with docetaxel produced objective responses in 21.1% and clinical benefits in 42.1% of the patients, respectively (1 complete response, 3 partial response, and 4 stable disease for at least 24 weeks, n = 19). Median time to progression was 148 days, and median overall survival was 579 days. The common adverse drug reactions were leukopenia, neutropenia, nail disorder, malaise, dysgeusia, alopecia, and edema.

CONCLUSIONS

TSU-68 in combination with docetaxel showed a promising antitumor response with manageable toxicity in patients with anthracycline-resistant metastatic breast cancer. Further studies are warranted in a different population of breast cancer or other solid cancers.

Human acidic <em>fibroblast</em> <em>growth</em> <em>factor</em> (haFGF) stimulates repair and regeneration of central and peripheral nerves after various injuries. However, it is unable to cross the blood-brain barrier (BBB). To produce a therapeutic haFGF with cell-permeable activity, we fused the haFGF(<em>19</em>-154) gene with Tat-PTD. After its construction by a single-step insertion of a polymerase chain reaction (PCR)-amplified coding sequence, the vector pTat-haFGF(<em>19</em>-154)-His was expressed in Escherichia coli BL21 (DE3) cells. The optimal expression level of the soluble fusion protein was up to 36.7% of the total cellular protein. The recombinant Tat-haFGF(<em>19</em>-154)-His was purified by a combination of Ni-NTA affinity, Sephadex G-25, and heparin affinity chromatography to 95% as detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The final yield was 171 mg/l culture. Purified Tat-haFGF(<em>19</em>-154)-His had distinct mitogenic activity in Balb/c 3T3 cells, as measured by methylthiazoletetrazolium (MTT) assay and its ED(50) was 3.931 x 10(-4) micromol/l. Tat-haFGF(<em>19</em>-154)-His protein intravenously injected at the dose of 10 mg/kg could be detected in the pallium and hippocampi.

BACKGROUND

This research elucidates the question of whether common and widespread dental procedures (DP) like root filling (RF) and the removal of wisdom teeth (WT) contribute to chronic inflammation in the jawbone. Dentists, in carrying out these DP, can set off defective wound healing in the jawbone in ignorance of its connection to inflammatory mediators and the possibility of it being a hidden cause of chronic systemic diseases (SYD).

METHODS

We examined samples of the jawbone for seven cytokines by multiplex analysis in three groups of jawbone areas. In order to clarify systemic interrelations, specimens from 16 patients were analyzed in areas of former surgery in the retromolar wisdom tooth area; specimens from 16 patients were analyzed in the jawbone, apically of teeth with RF; and specimens from <em>19</em> patients were of the healthy jawbone. Each of the retromolar and the apical jawbone samples showed clinically fatty degenerated and osteonecrotic medullary changes.

RESULTS

All fatty necrotic and osteolytic jawbone (FDOJ) samples showed regulated on activation, normal T-cell expressed and secreted (RANTES) and fibroblast growth factor (FGF)-2 as the only extremely overexpressed cytokines. FDOJ cohorts showed a 30-fold mean overexpression of RANTES and a 20-fold overexpressed level of FGF-2 when compared to healthy controls.

CONCLUSIONS

As RANTES is discussed in the literature as a possible contributor to inflammatory diseases, and though it might have oncogenic effects, we hypothesize that FDOJ in areas of improper and incomplete wound healing in the jawbone might act as hyperactivated signaling pathways, while serving as an unknown source of "silent inflammation". Because of the wide range of RANTES in immune diseases, treating FDOJ can cover many potential prediction or prognosis of individual outcomes.

BACKGROUND

Hepatocellular carcinoma (HCC), a primary neoplasm derived from hepatocytes, is the second leading cause of cancer mortality worldwide. Previous work has shown that <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>), an oncogenic driver, acts as a negative regulator of the therapeutic efficacy of the tyrosine kinase inhibitor sorafenib in HCC cells. The FGF<em>19</em>-mediated mechanism affecting sorafenib treatment, however, still remains to be resolved. Here, we hypothesize that the FGF<em>19</em>-FGFR4 axis may affect the effectiveness of sorafenib in the treatment of HCC.

METHODS

FGF<em>19</em> and FGFR4 cDNAs were cloned into a pcDNA3.1 vector and subsequently used for exogenous over-expression analyses. FGF<em>19</em> knockdown cells were generated using a lentiviral-mediated short hairpin RNA (shRNA) methodology and FGFR4 knockout cells were generated using a CRISPR-Cas9 methodology. FGFR4 activation in HCC cells was inhibited by BLU9931. The effects of exogenous gene over-expression, expression knockdown and knockout, as well as drug efficacies in HCC cells, were validated using Western blotting. HCC cell proliferation was assessed using a CellTiter 96® AQueous One Solution Cell Proliferation Assay, whereas NO levels were assessed using DAF-FM DA staining in conjunction with electrochemical biosensors.

RESULTS

We found that FGF<em>19</em>, when exogenously overexpressed, results in a reduced sorafenib-induced NO generation and a decreased proliferation of HCC cells. In contrast, we found that either FGF<em>19</em> silencing or knockout of its receptor FGFR4 sensitized HCC cells to sorafenib through the induction of NO generation. Concordantly, we found that inactivation of FGFR4 by BLU9931 enhanced the sensitivity of HCC cells to sorafenib.

CONCLUSIONS

From our data we conclude that the FGF<em>19</em>-FGFR4 axis may play a critical role in the effects elicited by sorafenib in HCC cells. Blocking the FGF<em>19</em>-FGFR4 axis may provide novel opportunities to improve the efficacy of sorafenib in the treatment of patients with HCC.

BACKGROUND

Acute kidney injury (AKI) increases the morbidity of critically ill children. Thus, it is necessary to identify better renal biomarkers to follow the outcome of these patients. This prospective case-control study explored the clinical value of a urinary biomarker profile comprised of neutrophil gelatinase lipocalin (uNGAL), fibroblast growth factor-2 (uFGF-2), and epidermal growth factor (uEGF) to follow these patients.

METHODS

Urine samples were collected from 21 healthy children, and 39 critically ill children (mean age 7.5 years ± 6.97 SD) admitted to a pediatric intensive care unit with sepsis or requiring extra corporeal membrane oxygenation (ECMO). uNGAL, uFGF-2, and uEGF levels were measured using ELISA kits during the first 24 h of admission to PICU, at peak of illness, and upon resolution of the critical illness.

RESULTS

On admission, the uNGAL and uFGF-2 levels were increased, and the uEGF levels were decreased, in critically ill children with AKI (n = 19) compared to those without AKI (n = 20), and healthy controls. A biomarker score using the combined cut-off values of uNGAL, uFGF-2, and uEGF (AUC = 0.90) showed the highest specificity to identify children with AKI, relative to each biomarker alone. uNGAL and uFGF-2 on admission showed high sensitivity and specificity to predict mortality (AUC = 0.82).

CONCLUSIONS

The biomarker profile comprised of uNGAL, uFGF-2, and uEGF increased the specificity to detect AKI in critically ill children, when compared to each biomarker used alone. uNGAL and uFGF-2 may also predict the risk of death. Further validation of these findings in a large sample size is warranted.

<AbstractText>The evaluation of patients with chronic watery diarrhea represents a diagnostic challenge for clinicians because organic causes, including inflammatory bowel disease, microscopic colitis, and chronic infection, must be differentiated from functional diarrhea and diarrhea-predominant irritable bowel syndrome. The purpose of this review is to summarize the available evidence on the usefulness of diagnostic tests in such patients.</AbstractText><AbstractText>We searched MEDLINE and EMBASE via OVID, from <em>19</em>78 until April 2017. We included diagnostic test accuracy studies reporting on the use of fecal and blood tests for the evaluation of adult patients with functional diarrhea, including irritable bowel syndrome. We assessed the risk of bias of included studies using a modified version of the Quality Assessment of Diagnostic Accuracy Studies II, and the certainty in the evidence using the GRADE (Grading of Recommendations Assessment, Development, and Evaluation) approach. We calculated pooled sensitivity and specificity, and the proportion of patients with true and false positive and negative results. We evaluated the following tests: erythrocyte sedimentation rate, C-reactive protein, fecal lactoferrin, fecal calprotectin, serologic tests for celiac disease, tests for bile acid diarrhea, the commercially available version of anti-cytolethal distending toxin B and anti-vinculin antibodies, and tests for Giardia infection. We did not evaluate breath tests for small intestinal bacterial over<em>growth</em>, as they are not part of a standard diarrhea workup.</AbstractText><p><div><b>RESULTS</b></div>Thirty-eight studies proved eligible to evaluate 1 or more of these tests. Erythrocyte sedimentation rate and C-reactive protein were similar at discriminating organic from functional disease, with sensitivity and specificity, respectively, of 0.54-0.78 and 0.46-0.95 for erythrocyte sedimentation rate and 0.73 and 0.78 for C-reactive protein. Among fecal tests, fecal calprotectin in a range of 50-60 μg/g (pooled sensitivity 0.81; 95% confidence interval [CI], 0.75-0.86; pooled specificity 0.87; 95% CI, 0.78-0.92) and fecal lactoferrin in a range of 4.0-7.25 μg/g (pooled sensitivity 0.79; 95% CI, 0.73-0.84; pooled specificity 0.93; 95%CI 0.63-0.99) presented the lowest proportion of false-negative results (low certainty in the evidence). Among tests for celiac disease, IgA tissue transglutaminase presented the best diagnostic test accuracy (sensitivity range, 0.79-0.99; specificity range, 0.90-0.99) with moderate certainty in the evidence. Among tests for bile acid diarrhea, the ⁷⁵selenium homotaurocholic acid test performed better than serum <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> and 7α-hydroxy-4-cholesten-3-one, but is not available in the United States. There was insufficient evidence to recommend serologic tests for irritable bowel syndrome at this time. There are several good diagnostic tests for Giardia infection.</p><AbstractText>Moderate to low certainty in the evidence indicates that available fecal and blood tests may play a role in the diagnostic workup of adult patients with functional diarrhea. At the moment, no tests are available to reliably rule in irritable bowel syndrome.</AbstractText>

BACKGROUND

Fibroblast growth factor-21 (FGF21) is an important metabolic regulator suggested to improve glucose metabolism and prevent dyslipidemia. An FGF21-resistant state that increases circulating FGF21 has been reported in obese patients. Although regular exercise prevents metabolic disease, the relationship of the fitness level to serum FGF21 level and body fat distribution in humans remains poorly understood.

OBJECTIVE

The objective of the study was to determine the relationship among the serum FGF21 concentration, cardiorespiratory fitness (CRF) level, and visceral fat area (VFA).

METHODS

Serum FGF21 was measured by an ELISA in 166 middle-aged and elderly Japanese men (aged 30-79 y) and 25 untrained and 21 endurance-trained young men (aged 19-29 y). CRF was assessed by measuring the peak oxygen uptake (VO2peak) and VFA by magnetic resonance imaging.

RESULTS

In the middle-aged and elderly subjects, the serum FGF21 level correlated with the VO2peak (r = -0.355, P < .001) and VFA (r = 0.487, P < .001). Stepwise multiple regression analysis showed VFA to be most strongly associated with the serum FGF21 level (β = .360, P < .001), and VO2peak was also an independent predictor of the serum FGF21 level (β = -.174, P = .019). Furthermore, the proportion of subjects with an FGF21 level below the limit of detection was significantly higher among the endurance-trained than among the untrained young men (71.4% vs 24.0%, P = .001), and the VO2peak and VFA were independently associated with an undetectable FGF21 level (P < .05).

CONCLUSIONS

CRF and VFA are key determinants of the circulating FGF21 concentration.

The purpose of the present study was to investigate whether human umbilical cord blood (UCB) as well as bone marrow (BM) can generate hepatocyte lineage cells in a simple culture condition. Mononuclear cells (MNCs) separated from UCB and BM were cultured in the presence of <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF)-1, FGF-2, stem cell <em>factor</em> (SCF), and hepatocyte <em>growth</em> <em>factor</em> (HGF). The cultured cells were analyzed for morphology and for the expression of mRNAs and/or proteins of hepatocyte lineage markers. Both the UCB and BM MNCs grown in the given culture condition yielded large, round cells that were adherent to the culture dishes. RT-PCR analysis showed that mRNAs of albumin (ALB), cytokeratin (CK)-18, and alpha-fetoprotein were expressed from day 7 in both the UCB- and BM-derived cells. Immunofluorescent staining showed that the large, round cells expressed not only ALB and CK-<em>19</em> but also proliferating cell nuclear antigen, implying the proliferative potential of hepatocyte lineage cells. Therefore, UCB as well as BM can give rise to hepatocyte lineage cells in the simple culture condition with HGF, SCF, FGF-1, and FGF-2. These cells may be one of the potential candidates of cell sources for the cytotherapy of hepatic disease, although it remains to be determined if the hepatocyte lineage cells are derived from plastic hematopoietic stem cells or from liver stem cells that reside in UCB or BM.