Effect of different alcohols on stratum corneum kallikrein 5 and phospholipase A2 together with epidermal keratinocytes and skin irritation.

OBJECTIVES: The aim of this exploratory study was to investigate the effect of ethanol, isopropanol and n-propanol on stratum corneum (SC) enzymes and keratinocytes in vitro together with their effects on skin condition and function. METHODS: Activities of kallikrein 5 (KLK5) and phospholipase A2 (PLA2) as well as keratinocyte metabolic activity, interleukin-1α (IL-1α) and tumor necrosis factor-α (TNF-α) were measured in vitro in the presence and absence of the different alcohols. We also measured transepidermal water loss (TEWL), skin capacitance, visual dryness and visual redness on the volar forearms of 25 Caucasian women following application of the alcohols 20 and 100 times per day over a period of 14 days in a clinical study. RESULTS: Reduced activities of KLK5 and PLA2 were observed in the presence of the alcohols. The greatest denaturing effect was always observed for n-propanol (P < 0.001), and in the case of PLA2, the effect of isopropanol was greater than ethanol (P < 0.001). Equally, ethanol had the mildest effects on keratinocyte metabolic activity and cytokine secretion (P < 0.001) and n-propanol always produced the most severe changes in normal and differentiated keratinocytes. These in vitro findings supported the clinical results where the major effects were on the induction of skin irritation (increased dropout rates) and ranked the intolerance of the different alcohols as follows: n-propanol > isopropanol > ethanol. At the high application frequencies, the effect of the different alcohols on transepidermal water loss (TEWL) and skin capacitance was similar, but at the low application frequencies, n-propanol had a significant effect on TEWL and capacitance values (P < 0.05). Equally, n-propanol and isopropanol produced significantly more skin redness at the low application frequencies. CONCLUSIONS: Clearly, isopropanol and n-propanol caused significant SC and keratinocyte perturbation in vitro together with damage to skin condition and function in vivo whereas ethanol did not. As a result, we show that ethanol-based sanitizers are better tolerated by skin, particularly in high-use settings, than other alcohols and should be the active ingredient of choice.

The Kenya cancer research and control stakeholder program: Evaluating a bilateral partnership to strengthen national cancer efforts.

Background: In response to a growing cancer burden and need for improved coordination among stakeholders in Kenya, the US National Cancer Institute and the Kenya Ministry of Health collaboratively hosted a stakeholder meeting in 2014 which identified four priority areas of need (research capacity building, pathology and cancer registries, cancer awareness and education, and health system strengthening) and developed corresponding action plans. Methods: Surveys were conducted with participants to collect input on the progress and impact of the 2014 stakeholder meeting. Findings: Of 69 eligible participants, 45 responded from academia, healthcare institutions, civil society, government, and international agencies. Of the four technical focus areas, three have continued to conduct working group meetings and two have conducted in-person meetings to review and update their respective action plans. Accomplishments linked to or enhanced by t meeting include: Kenyan and international support for expansion of population-based cancer registries, increased availability of prioritized diagnostic tests in selected regional referral hospitals, a greater focus on development of a national cancer research agenda, strategic planning for a community education strategy for cancer awareness, and improved coordination of partners through in-country technical assistance. Interpretation: The Stakeholder Program has successfully united individuals and organizations to improve cancer control planning in Kenya, and has enhanced existing efforts and programs across the country. This model of partners working in parallel on prioritized track activities has supported development of long term coordination of cancer research and control activities sustainable by the Kenyan government and Kenyan institutions.

Characterization of molecules mediating cell-cell communication in human cardiac valve interstitial cells.

Cell-cell interactions and adhesion determine cellular architectural organization, proliferation, signaling, differentiation, and death. We have identified the molecular components of different cell-cell junctions in human valve interstitial cells (ICs) both in situ and in culture. ICs were isolated, cultured, and phenotyped for cell surface and cytoplasmic markers by flow cytometry and immunocytochemistry. Western blotting was used to identify and quantify the molecular components of these cell-cell junctions in human valve ICs and compared with expression in smooth muscle and fibroblast cell types. N-cadherin and desmoglein were weakly detected on a low percentage of ICs, and the other classical cadherins were not detected. alpha- and beta-catenin, but not gamma-catenin, were expressed at equivalent levels by all valve ICs. Valve ICs did not express connexin-32 and -40; however, connexin-26 and -43 were equally expressed by a low percentage of ICs, demonstrating cell surface and cytoplasmic expression ,and connexin-45 was weakly expressed. The other cell types also expressed N-cadherin, alpha- and beta-catenin, desmoglein and connexin-43. The expression of these junctional molecules was predominantly by valve ICs on the inflow side of the valves. Human valve ICs have the ability to communicate with other valve ICs and mediate cell-cell adhesion via N-cadherin, connexin-26 and -43, and desmoglein. The junctions between valve ICs could support an interconnecting and coordinated cellular unit capable of controlling the functionality of the valve.

Reliable change assessment with practice effects in sport concussion research: a comment on Hinton-Bayre.

In his comments on our previous article, Hinton-Bayre advocates the use of the regression based approach in most cases of determining reliable change. This article comments on Hinton-Bayre's argument, discusses cases where the regression method might not be the preferred method, and presents adjustments that make the method more generally preferable.

Novel BTB/POZ domain zinc-finger protein, LRF, is a potential target of the LAZ-3/BCL-6 oncogene.

BTB/POZ-domain C2H2 zinc(Zn)-finger proteins are encoded by a subfamily of genes related to the Drosophila gap gene krüppel. To date, two such proteins, PLZF and LAZ-3/BCL-6, have been implicated in oncogenesis. We have now identified a new member of this gene subfamily which encodes a 62 kDa Zn-finger protein, termed LRF, with a BTB/POZ domain highly similar to that of PLZF. Both human and mouse LRF genes, which localized to syntenic chromosomal regions (19p13.3 and 10B5.3, respectively), were widely expressed in adult tissues and cell lines. At approximately 9.5-10.0 days of embryonic development, the mouse LRF gene was expressed in the limb buds, pharyngeal arches, tail bud, placenta and neural tube. The LRF protein associated in vivo with LAZ-3/BCL-6, but not with PLZF to which it was more related. Although the LRF, or LAZ-3/BCL-6, BTB/POZ domain could readily homodimerize, no heterodimerization was detected in vivo between the LRF and LAZ-3/BCL-6 BTB/POZ domains and interaction between full length LRF and LAZ-3/BCL-6 required the presence of both the BTB/POZ domain and Zn-fingers in each partner protein. As expected from the above results, LRF and LAZ-3/BCL-6 also colocalized with each other in the nucleus. Taken together, our findings suggest that BTB/ POZ-domain Zn-finger proteins may function as homo and heterodimeric complexes whose formation, and hence the resultant effect on transcription of their downstream target genes, is determined by the levels and expression domains of a given partner protein.

Heat shock treatment enhances graft cell survival in skeletal myoblast transplantation to the heart.

BACKGROUND: Graft survival after skeletal myoblast transplantation is affected by various pathological processes caused by environmental stress. Heat shock is known to afford protection of several aspects of cell metabolism and function. We hypothesized that prior heat shock treatment of graft cells would improve their survival after cell transplantation. METHODS AND RESULTS: L6 rat skeletal myoblasts expressing ss-galactosidase (ss-gal) were subjected to heat shock (42 degrees C, 1 hour). Increased expression of heat shock protein 72 was detected 24 hours later in the heat-shocked cells. After hypoxia-reoxygenation in vitro, lactate dehydrogenase leakage was significantly attenuated in the heat-shocked cells; in addition, the percentage of early apoptosis was lower in this group measured by flow cytometry with annexin V staining. For the in vivo study, 1 x 10(6) heat-shocked (hsCTx) or normal-cultured (CTx) myoblasts were infused into the explanted rat hearts through the coronary artery followed by heterotopic heart transplantation. ss-gal activity was significantly higher in the hsCTx group after cell transplantation, with an estimated 8 x 10(6) surviving cells per heart in the hsCTx group and 5 x 10(6) cells in the CTx group on day 28. Discrete loci of grafted cells were globally observed in the myocardium of the hsCTx and CTx groups, with a higher frequency in the hsCTx group. Surviving myoblasts occasionally differentiated into myotubes and had integrated with the native cardiomyocytes. CONCLUSIONS: Heat-shocked skeletal myoblasts demonstrated improved tolerance to hypoxia-reoxygenation insult in vitro and enhanced survival when grafted into the heart. Heat shock treatment could be useful in improving graft cell survival in cell transplantation.

Development of a novel method for cell transplantation through the coronary artery.

BACKGROUND: Cell transplantation is a promising strategy to treat end-stage heart failure. At present, a popular method to deliver cells into the heart is direct intramuscular injection. This method, however, may not be efficient in spreading cells globally into the myocardium. We have developed a novel method for cell transplantation using intracoronary infusion. METHODS AND RESULTS: An L6 rat skeletal muscle cell line expressing ss-galactosidase (ss-gal) was generated by gene transfection and clonal selection. These cells (10(6) in 1 mL medium) were infused into explanted rat hearts through the coronary artery, followed by heterotopic heart transplantation into the abdomen of recipients. Control hearts were infused with cell-free medium. According to ss-gal activity measurements, approximately 5 x 10(5) grafted cells per heart existed on day 3, increasing to 5 x 10(6) on day 28 in the cell-transplanted hearts. At day 28, discrete loci positively stained for ss-gal were observed throughout the cardiac layers of both left and right coronary territories. Some of them differentiated into ss-gal-positive multinucleated myotubes that aligned with the cardiac fiber axis and integrated into the native myocardium, whereas others formed colonies consisting of undifferentiated myoblasts. Connexin 43, a cardiac gap junction protein, was expressed between grafted cells and native cardiomyocytes. No reduction in cardiac function was observed in a Langendorff perfusion system. CONCLUSIONS: We have developed a unique method for efficient cell transplantation based on intracoronary infusion. This method, potentially applicable in the clinical setting during cardiac surgery, could be useful to globally supply cells to the heart.

A novel functional co-operation between MyoD, MEF2 and TRalpha1 is sufficient for the induction of GLUT4 gene transcription.

We report tripartite co-operation between MyoD, myocyte enhancer factor-2 (MEF2) and the thyroid hormone receptor (TRalpha1) that takes place in the context of an 82-bp muscle-specific enhancer in the rat insulin-responsive glucose transporter (GLUT4) gene that is active in both cardiac and skeletal muscle. In the L6E9 skeletal muscle cell line and in 10T1/2 fibroblasts, a powerful synergistic activation of the GLUT4 enhancer relied on the over-expression of MyoD, MEF2 and TRalpha1 and the integrity of their respective binding sites, and occurred when linked to either a heterologous promoter or in the context of the native GLUT4 promoter. In cardiac myocytes, enhancer activity was dependent on the binding sites for MEF2 and TRalpha1. Furthermore, we show that in 10T1/2 fibroblasts, the forced expression of MyoD, MEF2 and TRalpha1 induced the expression of the endogenous, otherwise silent, GLUT4 gene. In all, our results indicate a novel functional co-operation between these three factors which is required for full activation of GLUT4 transcription.

Identification of novel, cardiac-restricted transcription factors binding to a CACC-box within the human cardiac troponin I promoter.

OBJECTIVES: The expression of the human cardiac troponin I (hTnIc) gene is developmentally regulated and tissue-specific. In analysing the putative binding elements within the proximal promoter, a CACC-box sequence overlapping a consensus Sp1 element has been identified. The aim of this study was to characterise the factors binding to this element and to determine their importance in the transcriptional activity of the promoter. METHODS: A combination of supershift and competition electrophoretic mobility shift assays (EMSA) were used to identify the binding of factors to the overlapping CACC-box/Sp1 consensus element. The functional importance of this element was tested by transient transfection into primary neonatal rat cardiac myocytes in culture. RESULTS: At least four factors were able to interact with this region including the zinc finger proteins Sp1, Sp3 and two potentially novel factors. Whereas both Sp1 and Sp3 bound to the consensus Sp1 element, and to a lesser extent the CACC-box, two of the complexes required the intact CACC-box for binding. Site-directed mutagenesis of this region showed that the CACC-box is essential for hTnIc promoter-reporter activity. Further characterisation using EMSA indicated that the factors binding the hTnIc CACC-box are unlikely to be zinc finger proteins as they are insensitive to the addition of divalent cation chelating agents. They were also unable to bind to other known CACC-box elements. These factors are present in both human and rat cardiac muscle but absent from a number of cell lines including several derived from skeletal muscle. CONCLUSION: The human cardiac troponin I gene promoter requires an upstream CACC-box element for full activity. This element binds at least two complexes which represent novel, tissue-restricted DNA-binding activity present in the heart which we have named HCB1 and HCB2 for heart CACC-box binding factors.

Myocyte enhancer factor 2 (MEF2).

The Myocyte Enhancer Factor 2 (MEF2) proteins are transcription factors expressed during development of all three muscle lineages. Of the four mammalian mef2 genes, three (A, C and D) can be alternatively spliced, producing transcripts and proteins which may have significant functional differences. Specific binding sites for MEF2 proteins have been characterized in many striated muscle genes and MEF2 proteins can trans-activate gene expression both as homo- and heterodimers. Loss-of-function mutants in Drosophila indicate that MEF2 is an essential co-factor, but not a primary determinent, in the development of all three muscle lineages in the fly. Recent data suggest an interaction between the DNA-binding domains of mammalian MEF2 proteins and those of tissue-specific basic helix-loop-helix (bHLH) factors and thyroid hormone receptor alpha 1 (TR alpha 1) in the expression of target genes and the development of specific cell phenotypes. Understanding how MEF2 proteins function in the three mammalian muscle types may allow the development of therapeutic strategies for manipulating muscle growth and characteristics.

Origins of hyperphenylalaninemia in Israel.

Mutations and polymorphisms at the phenylalanine hydroxylase (PAH) gene were used to study the genetic diversity of the Jewish and Palestinian Arab populations in Israel. PAH mutations are responsible for a large variety of hyperphenylalaninemias (HPAs), ranging from the autosomal recessive disease phenylketonuria to various degrees of nonclinical HPA. Seventy-two Jewish and 36 Palestinian Arab families with various HPAs, containing 115 affected genotypes, were studied by haplotype analysis, screening for previously known PAH lesions and a search for novel mutations. Forty-one PAH haplotypes were observed in this sample. Four mutations previously identified in Europe (IVS10nt546, R261Q, R408W and R158Q) were found, and were associated with the same haplotypes as in Europe, indicating possible gene flow from European populations into the Jewish and Palestinian gene pools. Of particular interest is a PAH allele with the IVS10nt546 mutation and haplotype 6, that might have originated in Italy more than 3,000 years ago and spread during the expansion of the Roman Empire. These results, together with previous identification of three PAH mutations unique to Palestinian Arabs [IVSnt2, Edel(197-205) and R270S], indicate that the relatively high genetic diversity of the Jewish and Palestinian populations reflects, in addition to genetic events unique to these communities, some gene flow from neighboring and conquering populations.

Molecular cloning of the gene encoding the valyl-tRNA synthetase from Bacillus stearothermophilus.

The valS gene from the thermophile Bacillus stearothermophilus encoding the valyl-tRNA synthetase has been cloned on a 13.8-kb plasmid. The gene product and its kinetic properties are comparable with those of the native enzyme.

Molecular cloning of human cardiac troponin I using polymerase chain reaction.

We have used the polymerase chain reaction (PCR) to synthesise a cDNA encoding part of human cardiac troponin I. Amplification was achieved using fully degenerate sets of oligonucleotides corresponding to conserved regions of amino acid sequence identified in other troponin I isoforms. The cloned PCR fragment was subsequently used to isolate full-length cDNAs from a cardiac cDNA library. We describe the approach, as a general cloning strategy starting from limited amino-acid sequence data and report the cloning, and complete amino acid sequence of human cardiac troponin I. Analysis of human development using these clones demonstrates early expression of this gene in the heart.

Identification of the protein kinase C isoenzymes in human lung and airways smooth muscle at the protein and mRNA level.

The protein kinase C (PKC) isoenzymes expressed by human peripheral lung and tracheal smooth muscle resected from individuals undergoing heart-lung transplantation were identified at the protein and mRNA level. Western immunoblot analyses of human lung identified multiple PKC isoenzymes that were differentially distributed between the soluble and particulate fraction. Thus, PKC alpha, PKC betaII, PKC epsilon, and PKC zeta were recovered predominantly in the soluble fraction whereas the eta isoform was membrane-associated together with trace amounts of PKC alpha and PKC epsilon. PKC beta1-like immunoreactivity was occasionally seen although the intensity of the band was uniformly weak. Immunoreactive bands corresponding to PKCs gamma, delta, or theta were never detected. Reverse transcription-polymerase chain reaction (RT-PCR) of RNA extracted from human lung using oligonucleotide primer pairs that recognise unique sequences in each of the PKC genes amplified cDNA fragments that corresponded to the predicted sizes of PKC alpha, PKC betaI, PKC betaII, PKC epsilon, PKC zeta, and PKC eta (consistent with the expression of PKC isoenzyme protein) and, in addition, mRNA for PKC delta; PCR fragments of the expected size for the supposedly muscle-specific isoform, PKC theta, or the atypical isoenzyme, PKC lambda, were never obtained. The complement and distribution of PKC isoforms in human trachealis were similar, but not identical, to human lung. Thus, immunoreactive bands corresponding to the alpha, betaI, betaII, epsilon, and zeta isoenzymes of PKC were routinely labelled in the cytosolic fraction. In the particulate material PKC alpha, PKC epsilon, PKC alpha, PKC eta, and PKC mu were detected by immunoblotting. With the exception of PKC zeta, RT-PCR analyses confirmed the expression of the PKC isoforms detected at the protein level and, in addition, identified mRNA for PKC delta. Collectively, these data clearly demonstrate the expression of multiple PKC isoenzymes in human lung and tracheal smooth muscle, suggesting that they subserve diverse multifunctional roles in these tissues.

Possible heterogeneity in spondyloenchondrodysplasia: quadriparesis, basal ganglia calcifications, and chondrocyte inclusions.

We report on 6 patients with short stature and progressive enchondromatous-like changes of the vertebral bodies and the metaphyses of the long bones. Parental consanguinity was observed in 5 of 6 cases, supporting autosomal recessive inheritance. In spite of the similarity in radiographic changes and body proportions, genetic heterogeneity is suggested by the presence of CNS calcifications in 3 patients. Two of the latter had progressive quadriparesis. We tentatively classified these patients into 2 provisional types. An iliac crest biopsy in one of the patients with "type I" disease did not demonstrate enchondromatosis. Light and transmission electron microscopic studies demonstrated large cisterns and small inclusion bodies containing a flocculent material within the rough endoplasmic reticulum of the chondrocytes. Based on the histological and radiographic findings, we propose to classify these conditions among the spondylometaphyseal skeletal dysplasias.

Overexpression of connexin 43 in skeletal myoblasts: Relevance to cell transplantation to the heart.

OBJECTIVE: Skeletal myoblast transplantation is a promising strategy for treating end-stage heart failure. One potential problem in the development of functional, synchronously contracting grafts is the degree of intercellular communication between grafted myoblasts and host cardiomyocytes. Thus it is expected that enhancement of intercellular gap junction formation would result in improved efficiency of skeletal myoblast transplantation. In this study we investigated whether myoblasts overexpressing connexin 43, a major cardiac gap junction protein, would enhance this intercellular communication. METHODS AND RESULTS: L6 rat skeletal myoblast cell lines overexpressing connexin 43 were generated by means of gene transfection and clonal selection. Connexin 43 overexpression of these myoblasts, which continued both in undifferentiated and differentiated states (up to 17-fold greater protein level in comparison with control-transfected myoblasts, as measured with Western blotting), was observed on cell surfaces where gap junctions should exist. Both dye microinjection and scrape loading with fluorescent dyes showed enhancement in intercellular dye transfer between connexin 43-transfected myoblasts compared with that found in control-transfected cells. Morphologically, these myoblasts fused and differentiated into multinucleated myotubes more rapidly, demonstrating a higher level of cellular creatine kinase activity as a marker of myogenic differentiation throughout the culture period compared with that of control-transfected myoblasts. CONCLUSIONS: We have generated connexin 43-overexpressing skeletal myoblast cell lines that resulted in improved formation of functional intercellular gap junctions, which could be relevant to synchronous contraction of grafted myoblasts in the heart. In addition, these cells demonstrated more rapid differentiation, which would also be advantageous in a graft for transplantation to the heart.

Efficiency of a high-titer retroviral vector for gene transfer into skeletal myoblasts.

BACKGROUND: Genetic transformation of skeletal myoblasts for myocardial repair is dependent on an efficient gene transfer system that integrates the genes of interest into the genome of the target cell and its progeny. The aim of this investigation was to evaluate the use of a new retrovirally based gene transfer system for this purpose. METHODS: MFGnlslacZ retroviral vector, packaged in high-titer, split-genome packaging cell line (FLYA4) was used to transduce the skeletal myoblast cell line L6. L6 cells, cultured in 10% fetal calf serum, were transduced with the MFGnlslacZ vector by means of filtered supernatant from FLYA4 cells. Transduced L6 cells were divided into four groups. Group I cells were fixed as myoblasts 3 days after transduction. Group II cells were allowed to differentiate into myotubes. Group III cells were split every 3 days for 4 months. Group IV cells were split as in group III but then allowed to differentiate into myotubes. All samples were fixed and stained for beta-galactosidase activity. The effects on gene transfer of transforming growth factor-beta, insulin-like growth factor-I, and platelet-derived growth factor were determined by spectrophotometric assay of beta-galactosidase activity in cells transduced in the presence or absence of serum with 0 to 200 ng/ml of each growth factor. RESULTS: Morphometric analysis showed that 66.3% +/- 3% to 69.6% +/- 6% of cells in group I to IV expressed the lacZ reporter gene. In the presence of serum, transforming growth factor-beta significantly inhibited gene transfer, whereas insulin-like growth factor-I and platelet-derived growth factor significantly enhanced gene transfer. In absence of serum, however, only platelet-derived growth factor enhanced retrovirally mediated gene transfer into skeletal myoblasts. CONCLUSION: MFG retroviral vectors packaged in FLYA4 cells are efficient in gene transfer into skeletal myoblasts and result in transgenic expression that is maintained after repeated cell division, differentiation, or both. Platelet-derived growth factor enhances retrovirally mediated gene transfer into skeletal myoblasts.

Quality of life after laparoscopic adjustable gastric banding for severe obesity: postoperative and retrospective preoperative evaluations.

BACKGROUND: The aim of this study was to examine postoperative as well as retrospective preoperative evaluations of multiple dimensions of quality of life of patients with morbid obesity after laparascopic adjustable gastric banding (LAGB). METHODS: 12 to 38 months after LAGB, 74 consecutive patients (64 female, 10 male, mean age 36.6 years, age range 23-56) filled out the RAND-36 Health Survey questionnaire to evaluate their current postoperative as well as their past preoperative quality of life. RESULTS: Pre- to 1 year postoperative weight reduction (127.5 to 100.7 kg) and change of BMI (45.2 to 35.6 kg/m2) were highly significant (p<0.001). As compared to age reference groups, the preoperative quality of life was evaluated very poor (p<0.002), postoperative psychological and social quality of life were about normal (all p's >0.10), and postoperative physical functioning (p=0.04), vitality (p=0.01) and general health (p=0.03) were below normal. No differences were found between postoperative evaluations of patient groups with varying postoperative follow-up duration, but patients in the second year after surgery evaluated some aspects of their preoperative quality of life as poorer than patients in the third year after surgery. CONCLUSION: Postoperative psychosocial quality is at a level that may be expected to motivate patients to consolidate the surgically established weight reduction, but attention should be paid to the physical condition. Since the relative gain in quality of life as experienced by patients tends to be evaluated less with a longer duration of the postoperative interval, the risk of relapse may increase with passage of time.

Procedural learning of cognitive and motor skills in psychotic patients.

Two kinds of procedural learning, viz. learning of a sequence of simple motor responses and learning to solve a rather complex problem (Tower of Hanoi), as well as declarative learning (word list learning) were investigated in a group of psychotic inpatients (n = 67) and a control group of non-psychotic psychiatric inpatients (n = 19). Within the psychotic group, correlations of the task variables with positive and negative symptoms were explored. There was no difference between both groups in motor procedural learning. Psychotic patients were less efficient than controls in solving the Tower problem, but both groups again showed an equal amount of procedural learning. Consistent with the literature, however, a clear difference between both groups was found in declarative learning. The memory tasks did not correlate significantly with psychotic symptoms. These findings are interpreted as another indication that automatic information processing in psychotic patients is intact. The results are discussed with reference to neuropsychological research on procedural learning in neurological patients.

Expression of 5-hydroxytryptamine receptor subtype messenger RNA in interstitial cells from human heart valves.

BACKGROUND AND AIM OF THE STUDY: Severe heart valve disorder has been reported in patients receiving a combination of the anorectic drugs fenfluramine and phentermine. The exact molecular mechanisms involved remain unknown. Fenfluramine alters the serotonin level in the brain, while phentermine interferes with the pulmonary clearance of serotonin; these data suggest that serotonin levels affect regulation of valve function. The aim of the present study was to characterize the serotonin receptor (5-hydroxytryptamine) subtypes expressed in the interstitial cells of human heart valves. METHODS: Interstitial cells were isolated and cultured from the aortic, pulmonary, mitral and tricuspid valves of recipient hearts obtained during transplantation. Total RNA was extracted from cultured cells in order to determine gene expression by reverse transcription-polymerase chain reaction (RT-PCR) using 5-hydroxytryptamine (5-HT) subtype-specific primer pairs. RESULTS: The results show that: (i) 5-HT 1B and 1D receptor subtypes are expressed in all four heart valves. This is significant as the 1B and 1D receptor subfamilies are the target of the anti-migraine drug sumatriptan, and these receptors regulate cardiac function and movement; (ii) 5-HT 1A, 5-HT 1E and 5-HT 1F are not expressed in interstitial cells isolated from the valves. CONCLUSION: We conclude that preliminary evidence exists for the presence of distinct subsets of 5-HT receptors in human heart valves, indicating that interstitial cells of the valves potentially respond to serotonin levels.