OBJECTIVE

Short-term administration of delayed-release chenodeoxycholic acid to patients with irritable bowel syndrome with constipation (IBS-C) accelerates colonic transit and reduces symptoms. A preliminary study has shown that patients with IBS-C have reduced levels of bile acids (BAs) in feces and reduced synthesis of BA. We compared the levels of primary and secondary BAs in fecal samples collected over a 48-hour period from patients with IBS-C on a diet that contained 100 g fat per day, and compared them with levels in samples from healthy volunteers (controls). We also examined the relationship between overall colonic transit and biomarkers of BAs in patients with IBS-C.

METHODS

We performed a retrospective study of 45 patients with IBS-C and 184 controls. For controls, we estimated the 10th percentile of fasting serum levels of 7α-hydroxy-4-cholesten-3-one (C4, n = 184) and 48-hour fecal BAs (n = 46), and the 90th percentile of the fasting serum level of <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>, n = 50). Colonic transit was measured in patients using a validated scintigraphic method. Data from patients with IBS-C were analyzed using Spearman correlations to determine the relationships among levels of C4, FGF<em>19</em>, fecal BAs, and colonic transit.

RESULTS

Among the patients with IBS-C, 2 of 45 had low serum levels of C4, 4 of 43 had increased serum levels of FGF<em>19</em>, and 6 of 39 had low levels of BAs in feces collected over 48 hours. Patients with IBS-C had a significant increase in the proportions of fecal lithocholic acid compared with controls (P = .04), and a decrease in deoxycholic acid compared with controls (P = .03). In patients with IBS-C, there were inverse relationships between serum levels of C4 and FGF<em>19</em> and correlations among levels of 48-hour fecal BAs, colonic transit, and serum C4 and FGF<em>19</em>.

CONCLUSIONS

Approximately 15% of patients with IBS-C have reduced total BAs and level of deoxycholic acid in fecal samples collected over 48 hours on a 100 g fat diet. In these patients, lower levels of excretion of BAs into feces correlated with slower colonic transit.

Coronavirus disease 20<em>19</em> (COVID-<em>19</em>) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is presenting as a systemic disease associated with vascular inflammation and endothelial injury. Severe forms of SARS-CoV-2 infection induce acute respiratory distress syndrome (ARDS) and there is still an ongoing debate on whether COVID-<em>19</em> ARDS and its perfusion defect differs from ARDS induced by other causes. Beside pro-inflammatory cytokines (such as interleukin-1 β [IL-1β] or IL-6), several main pathological phenomena have been seen because of endothelial cell (EC) dysfunction: hypercoagulation reflected by fibrin degradation products called D-dimers, micro- and macrothrombosis and pathological angiogenesis. Direct endothelial infection by SARS-CoV-2 is not likely to occur and ACE-2 expression by EC is a matter of debate. Indeed, endothelial damage reported in severely ill patients with COVID-<em>19</em> could be more likely secondary to infection of neighboring cells and/or a consequence of inflammation. Endotheliopathy could give rise to hypercoagulation by alteration in the levels of different <em>factors</em> such as von Willebrand <em>factor</em>. Other than thrombotic events, pathological angiogenesis is among the recent findings. Overexpression of different proangiogenic <em>factors</em> such as vascular endothelial <em>growth</em> <em>factor</em> (VEGF), basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF-2) or placental <em>growth</em> <em>factors</em> (PlGF) have been found in plasma or lung biopsies of COVID-<em>19</em> patients. Finally, SARS-CoV-2 infection induces an emergency myelopoiesis associated to deregulated immunity and mobilization of endothelial progenitor cells, leading to features of acquired hematological malignancies or cardiovascular disease, which are discussed in this review. Altogether, this review will try to elucidate the pathophysiology of thrombotic complications, pathological angiogenesis and EC dysfunction, allowing better insight in new targets and antithrombotic protocols to better address vascular system dysfunction. Since treating SARS-CoV-2 infection and its potential long-term effects involves targeting the vascular compartment and/or mobilization of immature immune cells, we propose to define COVID-<em>19</em> and its complications as a systemic vascular acquired hemopathy.

Pharmacological doses of <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) 21 effectively normalize glucose, lipid and energy homeostasis in multiple animal models with many benefits translating to obese humans with type 2 diabetes. However, a role for FGF21 in the regulation of bile acid metabolism has not been reported. Herein, we demonstrate AAV-mediated FGF21 overexpression in mice increases liver expression of the key bile acid producing enzyme, Cyp7a1, resulting in an increased bile acid pool. Furthermore, in cholecystectomized mice, FGF21-mediated bile acid pool increase led to increased transit of bile acids into colon. We elucidate that the mechanism of FGF21 induced bile acid changes is mainly through antagonizing FGF15/<em>19</em> function on liver βKlotho/FGFR4 receptor complex; thus inhibiting FGF15/<em>19</em>-mediated suppression of Cyp7a1 expression. In conclusion, these data reveal a previously unidentified role for FGF21 on bile acid metabolism and may be relevant to understand the effects of FGF21 analogs in clinical studies.

BACKGROUND

βKlotho (βKL) is known to act as co-receptor for <em>fibroblast</em> <em>growth</em> <em>factor</em> receptor 4 (FGFR4) which is the main cognate receptor for <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>). Dysregulation of this FGF<em>19</em>/FGFR4/βKL signaling axis has been implicated in the pathogenesis of several cancers. However, its role in the pathogenesis of thyroid cancer has not been determined.

METHODS

The aim of this study was to assess FGF<em>19</em>, FGFR4 and βKL concentrations in a group of 36 patients with papillary thyroid cancer (PTC), 11 patients with follicular thyroid cancer (FTC), 9 patients with anaplastic thyroid cancer (ATC) and a group of <em>19</em> subjects with multinodular nontoxic goiter (MNG). The control group consisted of 20 healthy volunteers. Serum FGF<em>19</em>, FGFR4 and βKL concentrations were measured using specific ELISA methods.

RESULTS

Significantly lower concentrations of βKL and higher concentrations of FGF<em>19</em> were found in patients with PTC, FTC and ATC as compared with MNG group and controls. An elevation of FGFR4 serum concentration was observed in all thyroid cancer groups in comparison to MNG group and controls; however, in FTC group it was statistically insignificant. A positive correlation was found between βKL and FGFR4 concentrations in PTC patients. The levels of βKL, FGF<em>19</em> and FGFR4 did not differ significantly between MNG group and healthy controls.

CONCLUSIONS

Our results indicate that a disrupted FGF<em>19</em>/FGFR4/βKL signaling pathway may play a role in the development of thyroid cancers. However, further studies are needed to elucidate the molecular mechanism of the neoplastic transition of thyroid epithelial cells.

Myocardial fibrosis is often associated with cardiac hypertrophy; indeed, fibrosis is one of the most critical <em>factors</em> affecting prognosis. We aimed to identify the molecules involved in promoting fibrosis under hypertrophic stimuli. We previously established a rat model of cardiac hypertrophy by pulmonary artery banding, in which approximately half of the animals developed fibrosis in the right ventricle. Here, we first comprehensively analyzed mRNA expression in the right ventricle with or without fibrosis in pulmonary artery banding model rats by DNA microarray analysis (GSE141650 at NCBI GEO). The expression levels of <em>19</em> genes were up-regulated more than 1.5-fold in fibrotic hearts compared with non-fibrotic hearts. Among them, fibrosis <em>growth</em> <em>factor</em> (FGF) 23 showed one of the biggest increases in expression. Real-time PCR analysis also revealed that, among the FGF receptor (FGFR) family, FGFR1 was highly expressed in fibrotic hearts. We then found that FGF23 was expressed predominantly in cardiomyocytes, while FGFR1 was predominantly expressed in <em>fibroblasts</em> in the rat ventricle. Next, we added FGF23 and transforming <em>growth</em> <em>factor</em> (TGF)-β1 (10-50 ng/mL of each) to isolated <em>fibroblasts</em> from normal adult rat ventricles and cultured them for three days. While FGF23 itself did not directly affect the expression levels of any fibrosis-related mRNAs, FGF23 enhanced the effect of TGF-β1 on increasing the expression levels of α-smooth muscle actin (α-SMA) mRNA. This increase in xx-SMA mRNA levels due to the combination of TGF-β1 and FGF23 was attenuated by the inhibition of FGFR1 or the knockdown of FGFR1 in <em>fibroblasts</em>. Thus, FGF23 synergistically promoted the activation of <em>fibroblasts</em> with TGF-β1, transforming <em>fibroblasts</em> into myo<em>fibroblasts</em> via FGFR1. Thus, we identified FGF23 as a paracrine <em>factor</em> secreted from cardiomyocytes to promote cardiac fibrosis under conditions in which TGF-β1 is activated. FGF23 could be a possible target to prevent fibrosis following myocardial hypertrophy.

Recent findings have provided much insight into the mechanisms underlying long-term memory formation, and it is now known that long-term memory depends on the activation of a molecular cascade that culminates with structural changes in the brain. However, little is known about the signals that give rise to or regulate these structural changes. In this article we propose that <em>fibroblast</em> <em>growth</em> <em>factor</em>-2 (FGF2), a mitogen for several cell types, may be one of the molecular signals critically involved in the structural changes underlying long-term memory. If FGF2 is part of the signalling cascade involved in long-term memory, then increasing the activation of FGF2 should facilitate memory. In Experiments 1 and 2, we demonstrated that systemic injection of FGF2 (20ng/g of body weight) facilitated memory for contextual fear in 16, <em>19</em>, and 22 day old male Sprague Dawley rats. Experiment 3 demonstrated that the observed facilitation of memory was not due to FGF2 increasing rats' sensitivity to footshock. These results implicate FGF2 as a possible molecular signal in long-term memory, and further, illustrate a novel means of enhancing memory.

Variation in response to <em>growth</em> <em>factor</em> stimuli in myogenic satellite cell populations was investigated using a clonal-derived satellite cell culture system. Satellite cell clones were established from one muscle from one individual animal. One clone ("Early") which reached confluence on day <em>19</em> and one clone ("Late"), which reached confluence on day 29, were chosen for further examination. In previous studies, these two clones were found to differ in their <em>growth</em> rates in serum-containing medium. In the present study, the influence of <em>growth</em> <em>factors</em> on the proliferation of the two clones was compared in serum-free defined medium. Although basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (bFGF), insulin-like <em>growth</em> <em>factor</em>-I (IGF-I), insulin, and platelet-derived <em>growth</em> <em>factor</em>-BB (PDGF-BB) stimulated proliferation of both clones, the Early clone was more responsive to all <em>growth</em> <em>factors</em> tested than the Late clone (P < or = 0.05). The Early clone was also more responsive to the proliferative and differentiative depressing effects of administered transforming <em>growth</em> <em>factor</em>-beta (P < or = 0.05). Examination of properties of the PDGF, FGF, and IGF-I receptors on these two clones revealed no differences in either dissociation constants or receptor numbers (P>> or = 0.05). The results suggest that there is heterogeneity in satellite cell response to <em>growth</em> <em>factors</em>.

In quiescent <em>fibroblasts</em>, low-level transcription of the beta-actin gene occurs, and it is rapidly induced by <em>growth</em> <em>factors</em> in serum. Deletion analysis of the 5' upstream region of the human beta-actin gene using a CAT assay showed that the region between +765 and +783 downstream from the cap site, which had enhancer activity in <em>growing</em> NIH3T3 cells, also acted as a serum-response element in quiescent cells. The data suggest that the <em>19</em>-bp element acts positively as a serum response element. This sequence contains a CC(A/T)6GG sequence which is found in the promoter region of many cytoskeletal and muscle-specific actin genes.

A method for the in vitro proliferation of human bone marrow mesenchymal stem cells (MSCs) employing a medium not containing fetal calf serum (FCS) was developed for a regenerative medicine of cartilage using MSCs. Without using density-gradient centrifugation, the bone marrow aspirate was poured into a dish (6.0 \times 10(5) nucleated cells/cm(2)) with DMEM medium containing 10% serum (FCS or donor serum) and basic <em>fibroblast</em> <em>growth</em> <em>factor</em>, and incubated at 37 degrees C under a 5% CO(2) atmosphere. The density of adhesive cells incubated with the medium containing human serum and basic <em>fibroblast</em> <em>growth</em> <em>factor</em> (10 ng/ml) almost reached confluence at <em>19</em>d and was 1.4-2.7 times that in the medium containing only FCS. The density of cells incubated with the medium containing only human serum was 0.1-0.6 times that in the medium containing only FCS. The content of CD45(-) CD105(+) cells among the cells harvested after a <em>19</em>-d incubation in the medium containing human serum and basic <em>fibroblast</em> <em>growth</em> <em>factor</em> was higher than 90%. This high content and chondrogenic activity, which was confirmed by pellet cultivation and staining with Safranine O, were maintained even after further subcultivation in the medium to 17 population doubling levels. Consequently, this method might be applicable to in vitro proliferation of MSCs for the regeneration of cartilage.

Little progress has been made in identifying specific regulatory pathways that might be affected in cells by a mutationally activated p21ras when its expression does not lead to complete transformation. We wished to determine whether a normal, diploid human epithelial cell in which activation of ras had occurred could be identified in culture and, furthermore, whether expression of a mutant p21ras in such an otherwise normal cell would result in abnormal histogenic behavior in vivo. Thus, we introduced the v-Ha-ras gene into an early passage culture of normal human epidermal keratinocytes via a defective retrovirus. We examined these genetically engineered cells for changes in <em>growth</em> and differentiation, both in culture and in the epithelium formed when cultures were grafted to the skin of nude mice. We have found that keratinocytes expressing p21v-ras are independent of epidermal <em>growth</em> <em>factor</em> (EGF)--a <em>factor</em> which is normally essential for progressive colony <em>growth</em>, but that they are otherwise indistinguishable in culture from normal cells. v-ras keratinocytes also secrete a <em>factor</em> possessing some specific biological activities of members of the <em>fibroblast</em> <em>growth</em> <em>factor</em> (FGF) family, but which is distinct from acidic and basic FGF. In short-term dermal grafts the v-ras cells form a non-invasive and normally differentiating epidermis. However, the cells express elevated levels of keratin <em>19</em>, which is a characteristic of fetal epidermis and of premalignant lesions of some stratified squamous epithelia.

<em>Fibroblast</em> <em>growth</em> <em>factors</em> <em>19</em> and 21 (FGF<em>19</em> and FGF21) have been implicated, independently, in type 2 diabetes (T2D) but it is not known if their circulating levels correlate with each other or whether the associated hepatic signaling mechanisms that play a role in glucose metabolism are dysregulated in diabetes. We used a cross-sectional, case/control, experimental design involving Class III obese patients undergoing Roux-en-Y bariatric surgery (RYGB), and measured FGF<em>19</em> and FGF21 serum levels and hepatic gene expression (mRNA) in perioperative liver wedge biopsies. We found that T2D patients had lower FGF<em>19</em> and higher FGF21 serum levels. The latter was corroborated transcriptionally, whereby, FGF21, as well as CYP7A1, β-Klotho, FGFR4, HNF4α, and glycogen synthase, but not of SHP or FXR mRNA levels in liver biopsies were higher in T2D patients that did not remit diabetes after RYGB surgery, compared to T2D patients that remitted diabetes after RYGB surgery or did not have diabetes. In a Phenome-wide association analysis using 205 clinical variables, higher FGF21 serum levels were associated with higher glucose levels and various cardiometabolic disease phenotypes. When serum levels of FGF<em>19</em> were < 200 mg/mL and FGF21>> 500 mg/mL, 91% of patients had diabetes. These data suggest that FGF<em>19</em>/FGF21 circulating levels and hepatic gene expression of the associated signaling pathway are significantly dysregulated in type 2 diabetes.

Cirrhotic septa harbor vessels and inflammatory, fibrogenic, and ductular epithelial cells, collectively referred to as the ductular reaction (DR). Lack of the DR in the stromal compartment around hepatocellular carcinoma (HCC) has been documented; however, the relationship of epithelial keratin <em>19</em> (K<em>19</em>) structures to progression of intralesional carcinogenesis has not been explored. K<em>19</em> immunoreactivity in the stromal compartment around 176 nodules in cirrhotic explants was examined. Quantitative differences (P < 0.0001) were manifested in three distinct histologically identifiable patterns: "complex" around cirrhotic nodules (CN), "attenuated" around dysplastic nodules (DN), and "absent" around HCC. Markers of necrosis or apoptosis could not explain the perinodular K<em>19</em> epithelial loss; however, multicolor immunolabeling for K<em>19</em>, vimentin, E-Cadherin, SNAIL, and <em>fibroblast</em>-specific protein 1 (FSP-1) demonstrated discrepancies in immunophenotype and cytomorphologic features. Variability of cellular features was accompanied by an overall decrease in epithelial markers and significantly increased fractions of SNAIL- and FSP-1-positive cells in the DR around DN when compared with CN (P < 0.0001). Immunolabeling of transforming <em>growth</em> <em>factor</em>-β signaling components (TGFβR1, SMAD3, and pSMAD2/3) demonstrated increased percentages of pSMAD2/3 around DN when compared with CN (P < 0.0001). These findings collectively suggest marked alterations in cellular identity as an underlying mechanism for the reproducible extralesional K<em>19</em> pattern that parallels progressive stages of intranodular hepatocarcinogenesis. Paracrine signaling is proposed as a link that emphasizes the importance of the epithelial-stromal compartment in malignant progression of HCC in cirrhosis.

The <em>growth</em> and progression of most solid tumors depend on the initial transformation of the cancer cells and their response to stroma-associated signaling in the tumor microenvironment (1). Previously, research on the tumor microenvironment has focused primarily on tumor-stromal interactions (1-2). However, the tumor microenvironment also includes a variety of biophysical forces, whose effects remain poorly understood. These forces are biomechanical consequences of tumor <em>growth</em> that lead to changes in gene expression, cell division, differentiation and invasion(3). Matrix density (4), stiffness (5-6), and structure (6-7), interstitial fluid pressure (8), and interstitial fluid flow (8) are all altered during cancer progression. Interstitial fluid flow in particular is higher in tumors compared to normal tissues (8-10). The estimated interstitial fluid flow velocities were measured and found to be in the range of 0.1-3 μm s(-1), depending on tumor size and differentiation (9, 11). This is due to elevated interstitial fluid pressure caused by tumor-induced angiogenesis and increased vascular permeability (12). Interstitial fluid flow has been shown to increase invasion of cancer cells (13-14), vascular <em>fibroblasts</em> and smooth muscle cells (15). This invasion may be due to autologous chemotactic gradients created around cells in 3-D (16) or increased matrix metalloproteinase (MMP) expression (15), chemokine secretion and cell adhesion molecule expression (17). However, the mechanism by which cells sense fluid flow is not well understood. In addition to altering tumor cell behavior, interstitial fluid flow modulates the activity of other cells in the tumor microenvironment. It is associated with (a) driving differentiation of <em>fibroblasts</em> into tumor-promoting myo<em>fibroblasts</em> (18), (b) transporting of antigens and other soluble <em>factors</em> to lymph nodes (<em>19</em>), and (c) modulating lymphatic endothelial cell morphogenesis (20). The technique presented here imposes interstitial fluid flow on cells in vitro and quantifies its effects on invasion (Figure 1). This method has been published in multiple studies to measure the effects of fluid flow on stromal and cancer cell invasion (13-15, 17). By changing the matrix composition, cell type, and cell concentration, this method can be applied to other diseases and physiological systems to study the effects of interstitial flow on cellular processes such as invasion, differentiation, proliferation, and gene expression.

BACKGROUND

High-protein diets, which are popular for weight loss, contain large quantities of phosphorus. Phosphorus excess and consequent changes in phosphorus regulatory hormones are implicated in vascular calcification and cardiovascular disease.

OBJECTIVE

We tested the hypothesis that a moderate increase in dietary phosphorus during a high-protein diet leads to changes in phosphorus-responsive hormones.

METHODS

We conducted a post hoc analysis of a sequential dietary modification trial in <em>19</em> healthy volunteers in the general community.

METHODS

Participants received 2 weeks of a weight-maintaining, low-protein (15%) diet, followed by 2 weeks of an isocaloric, high-protein (30%) diet, followed by 12 weeks of an ad libitum high-protein (30%) diet.

METHODS

Using previously collected samples, plasma concentrations of fibroblast growth factor-23 (FGF-23), PTH, 1,25-dihydroxyvitamin D, and 24,25-dihydroxyvitamin D were measured at 8 time points to assess 24-hour variability and in 24-hour pooled samples to delineate changes at the end of each diet period.

RESULTS

Mean dietary phosphorus intake during each study period was 1556, 2071, and 1622 mg/d, respectively. Plasma concentrations of FGF-23 and vitamin D metabolites varied in a diurnal pattern; plasma PTH concentrations varied in a bimodal pattern. After changing from a low- to high-protein isocaloric diet, plasma FGF-23 concentrations decreased slightly (mean -4.48 pg/mL, 95% confidence interval 1.88-7.07). There were no other statistically significant changes in phosphorus regulatory hormones in response to diet modifications.

CONCLUSIONS

Among healthy people, an approximate 33% increase in dietary phosphorus after institution of a high-protein diet does not cause large changes in measured concentrations of phosphorus regulatory hormones.

OBJECTIVE

Prenatal tracheal occlusion or ligation (TL) has been proven to accelerate lung growth, but the mechanism of this is poorly understood. To increase understanding of the biological mechanisms involved in growth stimulation after TL in the fetal lung, we performed Global gene expression analysis using microarray technology.

METHODS

Sprague-Dawley rats underwent surgery on gestational day 19. After a small hysterotomy, the trachea was mobilized and tied. As controls, we used littermates to manipulated fetuses. On day 21, fetuses were removed and lungs harvested. Global gene expression analysis was performed using Affymetrix Platform and the RAE 230 set arrays (Affymetrix Inc, Santa Clara, Calif). For validation of microarray data, we performed real time polymerase chain reaction (PCR) of the most significant upregulated or downregulated genes, combined with immunohistochemical (IHC) analysis of lung sections.

RESULTS

In the group that underwent TL, several growth factors had an increased expression including connective tissue growth factor (CTGF), insulin-like growth factor 1 (IGF-1), and fibroblast growth factor 18 (FGF-18). Some of the genes that were downregulated in the group that underwent TL compared with controls were surfactant protein A (SP-A), apolipoprotein E (Apo-E), and phospholipase group II A2 (plg2a2). These results could be confirmed with real time PCR and IHC studies.

CONCLUSIONS

Tracheal occlusion or ligation is a well-documented stimulator of fetal lung growth, and the present study provides novel insights into the underlying molecular mechanisms, with increased expression of genes and proteins with growth factor activity. One of these growth factors, CTGF, has never been previously described in this model. Also, decreased levels of genes involved in surfactant metabolism were observed, providing molecular insights into the decreased surfactant production that is known to occur in TL. Increased understanding of the molecular mechanisms that control lung growth may be the key to develop novel therapeutic techniques to stimulate prenatal and/or postnatal lung growth.

<em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>) is a new member of the FGF family of <em>growth</em> <em>factors</em>. Here, we describe the localization of Fgf<em>19</em> mRNA in the developing chick retina and lens in stages from the Hamburger and Hamilton stage 15 (HH15) to postnatal day 30 (P30). Fgf<em>19</em> was expressed in a transient manner in postmitotic neuroblasts during the migration from the ventricular surface to their final location. Moreover, from HH31 (embryonic day 7, E7) on, a subset of lined up Fgf<em>19</em> expressing cells was distributed in the outer region of the presumptive INL. These cells were Pax6 immunoreactive horizontal cells. During the last third of embryogenesis, Fgf<em>19</em> expression in the retina was progressively down-regulated and was not detected at P30. Also, it was transiently expressed in the equatorial region of the lens.

Bile acid malabsorption occurs when there is impaired absorption of bile acids in the terminal ileum, so interrupting the normal enterohepatic circulation. The excess bile acids in the colon cause diarrhea, and treatment with bile acid sequestrants is beneficial. The condition can be diagnosed with difficulty by measuring fecal bile acids, or more easily by retention of selenohomocholyltaurine (SeHCAT), where this is available. Chronic diarrhea caused by primary bile acid diarrhea appears to be common, but is under-recognized where SeHCAT testing is not performed. Measuring excessive bile acid synthesis with 7α-hydroxy-4-cholesten-3-one may be an alternative means of diagnosis. It appears that there is no absorption defect in primary bile acid diarrhea but, instead, an overproduction of bile acids. <em>Fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>) inhibits hepatic bile acid synthesis. Defective production of FGF<em>19</em> from the ileum may be the cause of primary bile acid diarrhea.

Although the number of centenarians is <em>growing</em> worldwide, the potential <em>factors</em> influencing the aging process remain only partially elucidated. Researchers are increasingly focusing toward biomarkers as tools to shed more light on the pathophysiology of complex phenotypes, including the ability to reach successful aging, i.e., free of major chronic diseases. We therefore conducted a case-control study examining the potential associations of multiple candidate biomarkers in healthy centenarians and sex-matched healthy elderly controls. Using a case-control study of 81 centenarians (aged ≥ 100 years) selected based on the fact that they were disease-free and 46 healthy elderly controls (aged 70-80 years), serum levels of 15 different candidate biomarkers involved in the regulation of metabolism, angiogenesis, inflammation, and bone formation were measured. Of the 15 biomarkers tested, four molecules (chemerin, fetuin-A, and <em>fibroblast</em> <em>growth</em> <em>factors</em> [FGF] <em>19</em> and 21) were found to be independently associated with successful aging regardless of sex. Logistic regression analysis confirmed that chemerin, fetuin-A, FGF<em>19</em>, and FGF21 were independently associated with successful aging [predicted probability (PP) = 1 / [1 + 1 / exp (11.832 - 0.027 × (chemerin) - 0.009 × (fetuin-A) + 0.014 × (FGF<em>19</em>) - 0.007 × (FGF21)]. The area under the curve (AUC) of predicted probability values for the four-biomarker panel revealed that it can discriminate between centenarians and elderly controls with excellent accuracy (AUC>> 0.94, P < 0.001). Although preliminary in essence and limited by the low sample size and lack of replication in other independent cohorts, our data suggest an independent association between successful aging and serum chemerin, fetuin-A, FGF<em>19</em>, and FGF21, which may provide novel information on the mechanisms behind the human aging process. Whether the four-biomarker panel may predict successful aging deserves further scrutiny.

Adriamycin (doxorubicin hydrochloride), an effective chemotherapeutic drug, is also a potent inhibitor of wound healing. Conversely, certain polypeptide <em>growth</em> <em>factors</em> are capable of stimulating <em>fibroblasts</em> to secrete collagen, thus enhancing wound healing. The purpose of this study was to determine if interleukin-2 (IL-2), a T-cell <em>growth</em> <em>factor</em>, could reverse the wound healing deficit caused by Adriamycin. Adriamycin treatment caused a significant decrease in wound-breaking strength (P less than 0.005). IL-2 administration increased wound-breaking strength in Adriamycin-treated animals (2126 g vs 1549 g, P less than 0.005). In control animals, IL-2 did not increase wound-breaking strength significantly (2708 g vs 2608 g, P greater than 0.1). Histologically, wounds from Adriamycin-treated animals were less cellular, demonstrated less collagen in the dermis, and a lesser degree of capillary in<em>growth</em>. The number of <em>fibroblasts</em> in the dermal layer was increased in animals receiving IL-2. Control rats gained an average of 1.4% of their original body weight, while Adriamycin-treated rats lost an average of <em>19</em>% of their original body weight (P less than 0.0005). IL-2 administration did not influence weight loss or gain. Hematologically, animals receiving Adriamycin had lower hemoglobin and hematocrit values and higher platelet counts. There were no differences in total white blood cell counts; however, animals receiving Adriamycin showed a predominance of polymorphonuclear leukocytes and a relative decrease in lymphocytes. Animals receiving IL-2 demonstrated a significant eosinophilia. (1) Adriamycin impairs normal wound healing. (2) Interleukin-2 administration improves the wound healing impairment caused by Adriamycin. (3) Interleukin-2 appears to increase infiltration of inflammatory cells, <em>fibroblasts</em>, and capillaries into the wound, which may account for the observed increase in wound breaking strength.

The transforming <em>growth</em> <em>factor</em> beta (TGF-beta) plays an important role in constraining cellular proliferation, but it is also a potent inducer of programmed cell death or apoptosis. Here, we demonstrate that TGF-beta can have an opposite effect, acting as a survival <em>factor</em> to prevent c-Myc-induced cell death in Rat-1 <em>fibroblasts</em>. However, in marked contrast to TGF-beta, Smad2, which is a critical intracellular mediator of the TGF-beta signaling pathway, functions as an antagonist to induce increased cell death. The protective activity of TGF-beta was associated with the activation of c-Jun N-terminal Kinase (JNK) and was not linked to the ability of TGF-beta to promote cell cycle progression. Expression of dominant-interfering forms of various components of the JNK signaling pathway, including Rac1, Cdc42, mitogen-activated protein kinase kinase 4 (MKK4), and c-Jun, abolished TGF-beta-mediated cell survival. Furthermore, overexpression of the constitutively activated mutant RacL61F37A, which selectively stimulates JNK cascade but not G1 cell cycle progression or actin polymerization, was sufficient to prevent apoptosis induced by c-Myc. These findings describe a differential effect of two separated signaling pathways of TGF-beta and indicate for the first time that Smad2 can act as antagonist to suppress TGF-beta-dependent cell survival. Oncogene (2000) <em>19</em>, 1277 - 1287.

OBJECTIVE

Fibroblast growth factor 21 (FGF21) improves glucose and lipid metabolism, but high circulating levels are found in type 2 diabetes, suggesting FGF21 resistance. Serum FGF21 predicts incident diabetes, but its performance compared to established and emerging predictors is not known. We aimed to study the performance of FGF21 in diabetes prediction, relative to other adipokines and established risk factors including 2-h plasma glucose (2hG) during the oral glucose tolerance test (OGTT).

UNASSIGNED

We studied 1380 nondiabetic subjects from the Hong Kong Cardiovascular Risk Factor Prevalence Study using the second visit (2000-2004) as baseline when serum levels of FGF21 and other adipokines were measured. Glycaemic status was assessed by OGTT. Incident diabetes was defined as fasting glucose level (FG) ≥ 7 mmol/l or 2hG ≥ 11·1 mmol/l or use of antidiabetic agents, at subsequent visits.

RESULTS

A total of 123 participants developed diabetes over 9·0 years (median). On multivariable logistic regression analysis, FGF21 (P = 0·003), adipocyte fatty acid-binding protein (P = 0·003) and adiponectin (P = 0·035) were independent predictors of incident diabetes. FGF21 had the best change in log likelihood when added to a diabetes prediction model (DP) based on age, family history, smoking, hypertension, BMI, dyslipidaemia and FG. It also improved the area under ROC curve (AUROC) of diabetes prediction (DP) from 0·797 to 0·819 (P = 0·0072), rendering its performance comparable to the 'DP + 2hG' model (AUROC=0·838, P = 0·19).

CONCLUSIONS

As a biomarker for diabetes prediction, serum FGF21 appeared to be superior to other adipokines and, on its own, could be considered as an alternative to the OGTT.

BACKGROUND

Reabsorption of bile acids from the intestine by ileal bile acid transporter is pivotal for the enterohepatic circulation of BAs and sterol homoeostasis.

OBJECTIVE

To assess tolerability and study, bile acid metabolism in a phase 1 trial with the selective ileal bile acid transporter inhibitor A4250.

METHODS

A randomised double-blind, single-ascending dose (SAD) and multiple-ascending-dose study consisting of five cohorts comprising 40 individuals with a single administration of A4250 (0.1, 0.3, 1, 3, or 10 mg) or placebo and three cohorts comprising 24 individuals with a 1-week administration of A4250 (1 or 3 mg once daily or 1.5 mg twice daily) or placebo. For the multiple-ascending-dose study, bile acids were measured by HPLC-MS in plasma and faeces, and <em>fibroblast</em> <em>growth</em> <em>factor</em> <em>19</em> (FGF<em>19</em>) and 7α-hydroxy-4-cholesten-3-one (C4) were measured in plasma.

RESULTS

No serious adverse events occurred and all participants finished the trial per protocol. At the end of the multiple-ascending-dose study, plasma total bile acids and FGF<em>19</em> decreased by 47% and 76%, respectively, at 3 mg/day (P < 0.01), and by 15% and 16%, respectively, at 1.5 mg twice daily (P < 0.05). Plasma C4 and faecal bile acids increased at all dose regimens, by 555%, 664%, 292% and 338%, 421%, 420%, respectively (P < 0.01-0.05). The primary bile acids cholic and chenodeoxycholic acids constituted the majority of faecal bile acids in the A4250-treated groups.

CONCLUSIONS

A4250 is well tolerated. By blocking ileal bile acid transporter in the terminal ileum, it highly efficiently interrupts the enterohepatic circulation of BAs, and should be of benefit to patients with cholestatic liver diseases. Clinical Trial registration EudraCT 2013-001175-21.

Congenital GH insensitivity (Laron's syndrome, LS) is often associated with a dysfunctional GH receptor (GHR) causing complete insensitivity to GH and absent serum GH-binding protein (GHBP). However, a proportion of children with LS have normal GHBP levels. We have identified four girls from two families with this condition (height SD score, -3.4 to -6.8) and undertaken studies on 1) their GHR genes and 2) their GH responses in cultured skin <em>fibroblasts</em> to define the etiology of their GH insensitivities. No GHR gene mutations were identified in one family. In the other family, the affected siblings, an unaffected brother, and the father were heterozygous for a point mutation within exon 6 (D152H). In addition, use of intron 9 haplotypes to determine linkage to the GHR gene implied inheritance of different maternal GHR alleles in the two affected girls of the latter family. It is unlikely, therefore, that the D152H mutation alone could account for the LS phenotype. End points of GH action [DNA synthesis, insulin-like <em>growth</em> <em>factor</em>-binding protein-3 (IGFBP-3) messenger RNA (mRNA) and peptide production] in skin <em>fibroblast</em> cultures established from three of the LS subjects and four normal children were examined. Whereas normal <em>fibroblasts</em> incorporated [3H]thymidine dose dependently in response to 10-1000 ng/ml GH (increment at 1000 ng/ml, 77 +/- <em>19</em>%), LS <em>fibroblasts</em> failed to respond significantly above basal levels (P < 0.01). In normal <em>fibroblasts</em>, IGFBP-3 mRNA and peptide increased maximally at 48 h in response to 200 ng/ml GH, as determined by ribonuclease protection assay, Western ligand blotting, and RIA. In comparison, LS <em>fibroblasts</em> produced significantly less IGFBP-3 peptide than normal <em>fibroblasts</em> in response to GH, whereas IGFBP-3 mRNA failed to increase above basal levels. These studies have shown that 1) cultured human skin <em>fibroblasts</em> can be used to define the end points of GH action; 2) <em>fibroblast</em> cultures from the LS children show absent or reduced responses to GH; and 3) GH insensitivity in these children does not appear to be caused exclusively by GHR mutations, but is probably due to dysfunctional GHR signalling. Such patients may prove particularly important to elucidation of the key events in GH signaling.

OBJECTIVE

Reportedly, fibroblast growth factor receptor 3 (FGFR3) that regulates embryonic growth and development may function as an oncoprotein in certain malignancies. We aimed to investigate the biological significance of FGFR3 expression in invasive breast cancer.

METHODS

FGFR3 expression was investigated in 50 invasive breast cancer specimens by immunohistochemistry. The association between FGFR3 expression and clinicopathological/molecular parameters or prognosis was evaluated.

RESULTS

Weak FGFR3 expression was observed in myoepithelial cells, but not in duct epithelial cells, of the normal mammary ducts and lobules. FGFR3 expression in breast cancer cells was observed in 19 of 50 (38.0%) cases (9 weak positive and 10 strong positive). Besides the cytoplasm and cell membrane, nuclear staining was observed in 3 of 10 strong-positive cases. FGFR3 was further detected in non-neoplastic duct epithelial cells or duct papillomatosis in 5 strong-positive cases. No significant correlation was observed between FGFR3 expression and specific clinicopathological/molecular parameters. In contrast, FGFR3 expression was found to be significantly associated with overall survival in our cohort.

CONCLUSIONS

FGFR3 expression in invasive breast cancer was not found to be significantly associated with specific clinicopathological/molecular parameters, but might be used as a candidate marker for a poor prognosis.