bluuebunny/crossref_metadata_2025 huggingface dataset:
Long-term hydroxyurea therapy for infants with sickle cell anemia: the HUSOFT extension study
Authors: Hankins Jane S. , Ware Russell E. , Rogers Zora R. , Wynn Lynn W. , Lane Peter A. , Scott J. Paul , Wang Winfred C.
Published: June 2005

The long-term efficacy and toxicity of hydroxyurea for infants are undefined, and its role in preventing organ dysfunction is unknown. Short-term feasibility of hydroxyurea administration, toxicities, hematologic effects, and effect on spleen function in infants with sickle cell anemia (SCA) were reported (Hydroxyurea Safety and Organ Toxicity [HUSOFT] trial). These infants completing 2 years of hydroxyurea therapy (20 mg/kg/d) were offered study extension with dose escalation to 30 mg/kg/d. Patients were monitored with laboratory tests and biannual imaging studies. Hematologic indices were compared with predicted age-specific values and event rates compared with historic rates. All 21 subjects completing the original trial enrolled in the extension study: median age, 3.4 years old (range, 2.6 to 4.4 years); 12 females; 20 with Hb SS, 1 with Hb S/β0-thalassemia. Seventeen patients completed 4 years of hydroxyurea, and 11 completed 6 years. After 4 years, hydroxyurea was associated with increased hemoglobin concentration, percentage of fetal hemoglobin (Hb F), and mean corpuscular volume (MCV) and decreased reticulocytes, white blood cells (WBCs), and platelets (P < .01). Patients experienced 7.5 acute chest syndrome (ACS) events per 100 person-years, compared with 24.5 events per 100 person-years among historic controls (P = .001). Treated patients had better spleen function than expected and improved growth rates. Infants with SCA tolerate prolonged hydroxyurea therapy with sustained hematologic benefits, fewer ACS events, improved growth, and possibly preserved organ function.
Absence of inducible costimulator on alloreactive T cells reduces graft versus host disease and induces Th2 deviation
Authors: Hubbard Vanessa M. , Eng Jeffrey M. , Ramirez-Montagut Teresa , Tjoe Kartono H. , Muriglan Stephanie J. , Kochman Adam A. , Terwey Theis H. , Willis Lucy M. , Schiro Rafaella , Heller Glen , Murphy George F. , Liu Chen , Alpdogan Onder , van den Brink Marcel R. M.
Published: June 2005

Inducible costimulator (ICOS) is expressed on activated and memory T cells and is involved in the regulation of cytokine production. We studied the role of ICOS on alloreactive T cells in graft versus host disease (GVHD) and determined that ICOS expression was up-regulated on alloreactive T cells in recipients of an allogeneic hematopoietic stem cell transplantation (allo-HSCT) with GVHD. We compared ICOS-/- T cells with wild-type (WT) T cells in 2 GVHD models. In both models, recipients of ICOS-/- T cells demonstrated significantly less GVHD morbidity and mortality, which was associated with less intestinal and hepatic GVHD but increased cutaneous GVHD. In addition, recipients of ICOS-/- donor T cells displayed a slight decrease in graft versus leukemia (GVL) activity. Further analysis of alloreactive ICOS-/- T cells showed no defect in activation, proliferation, cytotoxicity, and target organ infiltration. Recipients of ICOS-/- T cells had decreased serum levels of interferon-γ (IFN-γ), while interleukin-4 (IL-4) and IL-10 levels were increased, suggesting that alloreactive ICOS-/- T cells are skewed toward T helper-2 (Th2) differentiation. These data suggest a novel role for ICOS in the regulation of Th1/Th2 development of activated T cells. In conclusion, alloreactive ICOS-/- donor T cells induce less GVHD due to a Th2 immune deviation while GVL activity is slightly diminished.
Triptolide, a constituent of immunosuppressive Chinese herbal medicine, is a potent suppressor of dendritic-cell maturation and trafficking
Authors: Chen Xin , Murakami Takaya , Oppenheim Joost J. , Howard O. M. Zack
Published: June 2005

Triptolide (TPT) is a chemically defined, potent immunosuppressive compound isolated from an anti-inflammatory Chinese herbal medicine. TPT has been reported to inhibit autoimmunity, allograft rejection, and graft-versus-host disease (GVHD), and its efficacy was previously attributed to the suppression of T cells. Since dendritic cells (DCs) play a major role in the initiation of T-cell–mediated immunity, we studied the effects of TPT on the phenotype, function, and migration of human monocyte–derived DCs. TPT treatment, over a pharmacologic concentration range, inhibited the lipopolysaccharide (LPS)–induced phenotypic changes, characteristic of mature DCs and the production of interleukin-12p70 (IL-12p70). Consequently, the allostimulatory functions of DCs were impaired by TPT treatment. Furthermore, the calcium mobilization and chemotactic responses of LPS-stimulated DCs to secondary lymphoid tissue chemokine (SLC)/CC chemokine ligand 21 (CCL21) were significantly lower in TPT-treated than untreated DCs, in association with lower chemokine receptor 7 (CCR7) and higher CCR5 expression. Egress of Langerhans cells (LCs) from explanted mouse skin in response to macrophage inflammatory protein-3β (MIP-3β)/CCL19 was arrested by TPT. In vivo administration of TPT markedly inhibited hapten (fluorescein isothiocyanate [FITC])–stimulated migration of mouse skin LCs to the draining lymph nodes. These data provide new insight into the mechanism of action of TPT and indicate that the inhibition of maturation and trafficking of DCs by TPT contributes to its immunosuppressive effects.
Dectin-1 activates Syk tyrosine kinase in a dynamic subset of macrophages for reactive oxygen production
Authors: Underhill David M. , Rossnagle Eddie , Lowell Clifford A. , Simmons Randi M.
Published: June 2005

Dectin-1 is a lectin receptor for β-glucan that is important for innate macrophage recognition of fungi and contributes to phagocytosis, reactive oxygen production, and induction of inflammatory cytokines. The mechanisms by which Dectin-1 mediates intracellular signaling are just beginning to be defined. Spleen tyrosine kinase (Syk) is a protein tyrosine kinase that is critical for adaptive immune responses where it mediates signaling through B-cell receptors, T-cell receptors, and Fc receptors. Here we report that Dectin-1 activates Syk in macrophages and is important for Dectin-1-stimulated reactive oxygen production, but not for phagocytosis. Syk activation is restricted to a subpopulation of macrophages that is in equilibrium with cells that cannot activate the pathway. The proportion of macrophages using this signaling pathway can be modulated by cytokine treatment. Thus, Dectin-1 signaling reveals dynamic macrophage heterogeneity in inflammatory activation potential. (Blood. 2005;106:2543-2550)
Altered body iron distribution and microcytosis in mice deficient in iron regulatory protein 2 (IRP2)
Authors: Galy Bruno , Ferring Dunja , Minana Belen , Bell Oliver , Janser Heinz G. , Muckenthaler Martina , Schümann Klaus , Hentze Matthias W.
Published: June 2005

Iron regulatory protein 2 (IRP2)-deficient mice have been reported to suffer from late-onset neurodegeneration by an unknown mechanism. We report that young adult Irp2-/- mice display signs of iron mismanagement within the central iron recycling pathway in the mammalian body, the liver-bone marrow-spleen axis, with altered body iron distribution and compromised hematopoiesis. In comparison with wild-type littermates, Irp2-/- mice are mildly microcytic with reduced serum hemoglobin levels and hematocrit. Serum iron and transferrin saturation are unchanged, and hence microcytosis is not due to an overt decrease in systemic iron availability. The liver and duodenum are iron loaded, while the spleen is iron deficient, associated with a reduced expression of the iron exporter ferroportin. A reduction in transferrin receptor 1 (TfR1) mRNA levels in the bone marrow of Irp2-/- mice can plausibly explain the microcytosis by an intrinsic defect in erythropoiesis due to a failure to adequately protect TfR1 mRNA against degradation. This study links a classic regulator of cellular iron metabolism to systemic iron homeostasis and erythropoietic TfR1 expression. Furthermore, this work uncovers aspects of mammalian iron metabolism that can or cannot be compensated for by the expression of IRP1. (Blood. 2005;106: 2580-2589)
Activated platelets induce Weibel-Palade–body secretion and leukocyte rolling in vivo: role of P-selectin
Authors: Dole Vandana S. , Bergmeier Wolfgang , Mitchell Heather A. , Eichenberger Sarah C. , Wagner Denisa D.
Published: June 2005

The presence of activated platelets and platelet-leukocyte aggregates in the circulation accompanies major surgical procedures and occurs in several chronic diseases. Recent findings that activated platelets contribute to the inflammatory disease atherosclerosis made us address the question whether activated platelets stimulate normal healthy endothelium. Infusion of activated platelets into young mice led to the formation of transient platelet-leukocyte aggregates and resulted in a several-fold systemic increase in leukocyte rolling 2 to 4 hours after infusion. Rolling returned to baseline levels 7 hours after infusion. Infusion of activated P-selectin-/- platelets did not induce leukocyte rolling, indicating that platelet P-selectin was involved in the endothelial activation. The endothelial activation did not require platelet CD40L. Leukocyte rolling was mediated solely by the interaction of endothelial P-selectin and leukocyte P-selectin glycoprotein ligand 1 (PSGL-1). Endothelial P-selectin is stored with von Willebrand factor (VWF) in Weibel-Palade bodies. The release of Weibel-Palade bodies on infusion of activated platelets was indicated by both elevation of plasma VWF levels and by an increase in the in vivo staining of endothelial P-selectin. We conclude that the presence of activated platelets in circulation promotes acute inflammation by stimulating secretion of Weibel-Palade bodies and P-selectin–mediated leukocyte rolling.
Feature selection and classification for microarray data analysis: Evolutionary methods for identifying predictive genes
Authors: Jirapech-Umpai Thanyaluk , Aitken Stuart
Published: June 2005

Background

In the clinical context, samples assayed by microarray are often classified by cell line or tumour type and it is of interest to discover a set of genes that can be used as class predictors. The leukemia dataset of Golub et al. [1] and the NCI60 dataset of Ross et al. [2] present multiclass classification problems where three tumour types and nine cell lines respectively must be identified. We apply an evolutionary algorithm to identify the near-optimal set of predictive genes that classify the data. We also examine the initial gene selection step whereby the most informative genes are selected from the genes assayed.

Results

In the absence of feature selection, classification accuracy on the training data is typically good, but not replicated on the testing data. Gene selection using the RankGene software [3] is shown to significantly improve performance on the testing data. Further, we show that the choice of feature selection criteria can have a significant effect on accuracy. The evolutionary algorithm is shown to perform stably across the space of possible parameter settings – indicating the robustness of the approach. We assess performance using a low variance estimation technique, and present an analysis of the genes most often selected as predictors.

Conclusion

The computational methods we have developed perform robustly and accurately, and yield results in accord with clinical knowledge: A Z-score analysis of the genes most frequently selected identifies genes known to discriminate AML and Pre-T ALL leukemia. This study also confirms that significantly different sets of genes are found to be most discriminatory as the sample classes are refined.
Genome-wide structural and evolutionary analysis of the P450 monooxygenase genes (P450ome) in the white rot fungus Phanerochaete chrysosporium : Evidence for gene duplications and extensive gene clustering
Authors: Doddapaneni Harshavardhan , Chakraborty Ranajit , Yadav Jagjit S
Published: June 2005

Background

Phanerochaete chrysosporium, the model white rot basidiomycetous fungus, has the extraordinary ability to mineralize (to CO2) lignin and detoxify a variety of chemical pollutants. Its cytochrome P450 monooxygenases have recently been implied in several of these biotransformations. Our initial P450 cloning efforts in P. chrysosporium and its subsequent whole genome sequencing have revealed an extraordinary P450 repertoire ("P450ome") containing at least 150 P450 genes with yet unknown function. In order to understand the functional diversity and the evolutionary mechanisms and significance of these hemeproteins, here we report a genome-wide structural and evolutionary analysis of the P450ome of this fungus.

Results

Our analysis showed that P. chrysosporium P450ome could be classified into 12 families and 23 sub-families and is characterized by the presence of multigene families. A genome-level structural analysis revealed 16 organizationally homogeneous and heterogeneous clusters of tandem P450 genes. Analysis of our cloned cDNAs revealed structurally conserved characteristics (intron numbers and locations, and functional domains) among members of the two representative multigene P450 families CYP63 and CYP505 (P450foxy). Considering the unusually complex structural features of the P450 genes in this genome, including microexons (2–10 aa) and frequent small introns (45–55 bp), alternative splicing, as experimentally observed for CYP63, may be a more widespread event in the P450ome of this fungus. Clan-level phylogenetic comparison revealed that P. chrysosporium P450 families fall under 11 fungal clans and the majority of these multigene families appear to have evolved locally in this genome from their respective progenitor genes, as a result of extensive gene duplications and rearrangements.

Conclusion

P. chrysosporium P450ome, the largest known todate among fungi, is characterized by tandem gene clusters and multigene families. This enormous P450 gene diversity has evolved by extensive gene duplications and intragenomic recombinations of the progenitor genes presumably to meet the exceptionally high metabolic demand of this biodegradative group of basidiomycetous fungi in ecological niches. In this context, alternative splicing appears to further contribute to the evolution of functional diversity of the P450ome in this fungus. The evolved P450 diversity is consistent with the known vast biotransformation potential of P. chrysosporium. The presented analysis will help design future P450 functional studies to understand the underlying mechanisms of secondary metabolism and oxidative biotransformation pathways in this model white rot fungus.
Haploinsufficiency for BRCA1is associated with normal levels of DNA nucleotide excision repair in breast tissue and blood lymphocytes
Authors: Latimer Jean J , Rubinstein Wendy S , Johnson Jennifer M , Kanbour-Shakir Amal , Vogel Victor G , Grant Stephen G
Published: June 2005

Background

Screening mammography has had a positive impact on breast cancer mortality but cannot detect all breast tumors. In a small study, we confirmed that low power magnetic resonance imaging (MRI) could identify mammographically undetectable tumors by applying it to a high risk population. Tumors detected by this new technology could have unique etiologies and/or presentations, and may represent an increasing proportion of clinical practice as new screening methods are validated and applied. A very important aspect of this etiology is genomic instability, which is associated with the loss of activity of the breast cancer-predisposing genesBRCA1andBRCA2. In sporadic breast cancer, however, there is evidence for the involvement of a different pathway of DNA repair, nucleotide excision repair (NER), which remediates lesions that cause a distortion of the DNA helix, including DNA cross-links.

Case presentation

We describe a breast cancer patient with a mammographically undetectable stage I tumor identified in our MRI screening study. She was originally considered to be at high risk due to the familial occurrence of breast and other types of cancer, and after diagnosis was confirmed as a carrier of a Q1200X mutation in theBRCA1gene. In vitro analysis of her normal breast tissue showed no differences in growth rate or differentiation potential from disease-free controls. Analysis of cultured blood lymphocyte and breast epithelial cell samples with the unscheduled DNA synthesis (UDS) assay revealed no deficiency in NER.

Conclusion

As new breast cancer screening methods become available and cost effective, patients such as this one will constitute an increasing proportion of the incident population, so it is important to determine whether they differ from current patients in any clinically important ways. Despite her status as aBRCA1mutation carrier, and her mammographically dense breast tissue, we did not find increased cell proliferation or deficient differentiation potential in breast epithelial cells from this patient which might have contributed to her cancer susceptibility. Although NER deficiency has been demonstrated repeatedly in blood samples from sporadic breast cancer patients, analysis of blood cultured lymphocytes and breast epithelial cells for this patient proves definitively that heterozygosity for inactivation ofBRCA1does not intrinsically confer this type of genetic instability. These data suggest that the mechanism of genomic instability driving the carcinogenic process may be fundamentally different in hereditary and sporadic breast cancer, resulting in different genotoxic susceptibilities, oncogene mutations, and a different molecular pathogenesis.
Seroprevalence and risk factors for toxoplasmainfection among pregnant women in Aydin province, Turkey
Authors: Ertug Sema , Okyay Pinar , Turkmen Munevver , Yuksel Hasan
Published: June 2005

Background

The aims of the present study were to determine the prevalence of toxoplasmosis in pregnant women at first trimester of their pregnancy and to follow up the seroconversion for next two trimesters, and to identify the risk factors and possible contamination routes in Aydin province, Turkey.

Method

The sample size was calculated as 423 on a prevalence of 50%, d=0.05 at a confidence level of 95% with 10% addition. It was a cross-sectional study with multistage sampling. After a questionnaire applied to the pregnant women, anti-Toxoplasma IgG antibodies were studied with ELISA and IFA, values in conflict with DA test, where IgM antibodies were studied with ELISA and for borderline or positive values of IgM avidity test was used.

Results

The mean age of 389 (92.9%) of pregnant women in the study was 24.28+/-4.56 years, the seroprevalence of anti-Toxoplasma IgG antibodies for toxoplasmosis was 30.1%. Seroprevalence was increased with age (p=0.001) and with drinking water consumption other than bottled water (p=0.042). No significant relations were observed between anti-Toxoplasma IgG antibodies and education level, being native or migrant, abortion history, consumption of meat, vegetable and milk/milk products, personal or kitchen hygiene habits, cat owning at home of the pregnant women. No IgM antibody was detected.

Conclusion

One of every three pregnant women in Aydin was at risk of toxoplasmosis at the first trimester of their pregnancy. Increased seroprevalance with age was a predictable result because of increasing time of exposure. Increased seroprevalence with consumption of municipal and uncontrolled water (well/spring water) supplies was similar with latest epidemiological findings.
Infrared thermography fails to visualize stimulation-induced meridian-like structures
Authors: Litscher Gerhard
Published: June 2005

Background

According to Traditional Chinese Medicine (TCM) the vital energy flows through a system of channels also called meridians. Generally accepted proof for meridians cannot be considered as being given. Goal of this study was to examine whether possible stimulation-induced meridian-like structures, as recently described by other authors, can be visualized and objectified simultaneously at different infrared wavelength ranges.

Methods

The study analyses evidence for the existence of acupuncture-specific, meridian-like artifacts in 6 healthy volunteers (mean age ± SD 28.7 ± 3.7 years; range 25 – 35 years). Two infrared cameras at different wavelength ranges were used for thermographic control of possible stimulation effects (moxibustion-cigar, infrared warmth stimulation, needle and laserneedle stimulation). In addition to thermography, temperature and microcirculatory parameters were registered at a selected point using laser-Doppler flowmetry.

Results and Conclusion

After moxibustion (or infrared light stimulation) of the body at 2 – 5 μm and 7.5 – 13 μm ranges, different structures appear on thermographic images of the human body which are technical artifacts and which are not identical to what are known as meridians in all textbooks of TCM. Further scientific studies are required regarding the possible visualization of meridians.
Acquisition of rifabutin resistance by a rifampicin resistant mutant of Mycobacterium tuberculosis involves an unusual spectrum of mutations and elevated frequency
Authors: Anthony Richard M , Schuitema Anja RJ , Bergval Indra L , Brown Tim J , Oskam Linda , Klatser Paul R
Published: June 2005

Background

Mutations in a small region of therpoBgene are responsible for most rifamycin resistance inMycobacterium tuberculosis. In this study we have sequentially generated resistant strains to first rifampicin and then rifabutin. Portions of therpoBgene were sequenced from 131 randomly selected mutants. Second round selection resulted in a changed frequency of specific mutations.

Methods

Mycobacterium tuberculosis(strain Mtb72) rifamycin resistant mutants were selectedin vitrowith either rifampicin or rifabutin. One mutant R190 (rpoBS522L) selected with rifampicin had a rifampicin MIC of 32 μg/ml but remained sensitive to rifabutin (MIC<0.8 μg/ml). This mutant was subjected to a second round of selection with rifabutin.

Results

All 105 first round resistant mutants derived from the parent strain (Mtb72) screened acquired mutations within the 81 bprpoBhotspot. When the rifampicin resistant but rifabutin sensitive S522L mutant was subjected to a second round of selection, single additionalrpoBmutations were identified in 24 (92%) of 26 second round mutants studied, but 14 (54%) of these strains contained mutations outside the 81 bp hotspot (codons 144, 146, 148, 505). Additionally, spontaneous rifabutin resistant mutants were produced at >10 times the frequency by the S522L mutant than the parent strain.

Conclusion

First round selection of mutation S522L with rifampicin increased the frequency and changed the spectrum of mutations identified after selection with rifabutin.
Long term remission of metastatic placental site trophoblastic tumor (PSTT): Case report and review of literature
Authors: Behtash Nadereh , Ghaemmaghami Fatemeh , Hasanzadeh Malihe
Published: June 2005

Background

Placental site trophoblastic tumor (PSTT) is a rare and unique form of gestational trophoblastic disease (GTD). This tumor represents a neoplastic transformation of intermediate trophoblastic cells. We document a case of long term remission in a patient with metastatic PSTT.

Case presentaion

A 27-year-old patient with metastatic PSTT was treated with combination therapy (chemotherapy and surgery). Patient is alive after 10 years without any evidence of recurrence. Literature on PSTT was searched using Medline and cross references, and pertinent articles were reviewed.

Conclusion

With surgery and chemotherapy it is possible to achieve long-term remission in metastatic PSTT. Only a handful of previously reported cases with prolonged remission had been treated with the described combined chemotherapy and surgical approach. We suggest that this approach may be recommended for metastatic PSTT.
Second language acquisition and first language loss in adult early bilinguals: exploring some differences and similarities
Authors: Montrul Silvina
Published: June 2005

This study compares the linguistic knowledge of adult second language (L2) learners, who learned the L2 after puberty, with the potentially ‘eroded’ first language (L1) grammars of adult early bilinguals who were exposed to the target language since birth and learned the other language simultaneously, or early in childhood (before age 5). I make two main claims: (1) that the L1 grammar of bilinguals at a given stabilized state (probably endstate) resembles the incomplete (either developing or stabilized) grammars typical of intermediate and advanced stages in L2 acquisition; and (2) that despite similar patterns of performance, when language proficiency is factored in, early bilinguals are better than the L2 learners, probably due to exposure to primary linguistic input early in childhood.

I offer empirical evidence from an experimental study testing knowledge of the syntax and semantics of unaccusativity in Spanish, conducted with English-speaking L2 learners and English-dominant Spanish heritage speakers living in the USA. I consider recent treatments of unaccusativity and language attrition within the generative framework (Sorace, 1999; 2000a; 2000b), that offer a unifying account of the formal parallels observed between these two populations I discuss how input, use and age may explain differences and similarities in the linguistic attainment of the two groups.
Characterization of the Mouse and Rat Relaxin Receptors
Authors: SCOTT D J , LAYFIELD S , RIESEWIJK A , MORITA H , TREGEAR G W , BATHGATE R A D
Published: June 2005

Abstract: Rodent models have been used for many years to probe the actions of relaxin. Identification of the orthologs of human leucine‐rich repeat‐containing g‐protein‐coupled receptor 7 (LGR7), the relaxin receptor, in mouse and rat will enable characterization of the response of LGR7 to relaxin in these species. Partial LGR7 homologous sequences from mouse and rat were discovered in the Celera and NCBI gene databases, amplified, cloned, and sequenced. At the protein level, mouse and rat LGR7 are 85.2% and 85.7% identical to human LGR7. Mouse and rat LGR7 were able to bind to and be activated by relaxin ligands.
LGR7‐Truncate Is a Splice Variant of the Relaxin Receptor LGR7 and Is a Relaxin Antagonist in Vitro
Authors: SCOTT D J , TREGEAR G W , BATHGATE R A D
Published: June 2005

Abstract: The relaxin receptor (LGR7) and the insulin‐like peptide 3 (INSL3) receptor (LGR8) are unique LGR family members in possessing a single, functionally important amino terminal LDL‐A module.1 Mouse and rat cDNA was screened for LGR7 and LGR7 splice variant expression. A uterus‐specific exon 4 deleted variant was identified and named LGR7‐Truncate. Deletion of exon 4 results in a premature stop codon and a transcript that putatively encodes a secreted protein containing LGR7's LDL‐A module. Expression of LGR7‐Truncate with LGR7 in HEK‐293T cells resulted in decreased relaxin‐induced signaling of LGR7. LGR7‐Truncate is potentially an endogenous regulator of LGR7 signaling.
Studies on Soluble Ectodomain Proteins of Relaxin (LGR7) and Insulin 3 (LGR8) Receptors
Authors: YAN YAN , CAI JIN , FU PING , LAYFIELD SHARON , FERRARO TANIA , KUMAGAI JIN , SUDO SATOKO , TANG JIAN‐GUO , GIANNAKIS ELENI , TREGEAR GEOFFREY W. , WADE JOHN D. , BATHGATE ROSS A.D.
Published: June 2005

Abstract: The ectodomains of both the relaxin (LGR7) and the INSL3 (LGR8) receptors can be expressed on the cell surface using only a single transmembrane domain. These membrane‐anchored proteins retain the ability to bind relaxin and can be cleaved from the cell surface. The subsequent LGR7 protein, 7BP, binds relaxin and can act as a functional relaxin antagonist. By contrast, the equivalent LGR8 protein 8BP does not bind relaxin or antagonize LGR8 activity. The 7BP protein has been successfully immobilized onto chemically derivatized surfaces for the capture of relaxin peptides and subsequent identification via SELDI‐MS analysis.
Receptors for Relaxin Family Peptides
Authors: BATHGATE ROSS A. , IVELL RICHARD , SANBORN BARBARA M. , SHERWOOD O DAVID , SUMMERS ROGER J.
Published: June 2005

Abstract: Recent studies have identified four receptors that are the physiological targets for relaxin family peptides. All are class I (rhodopsin like) G‐protein‐coupled receptors with LGR7 (RXFP1) and LGR8 (RXFP2) being type C leucine‐rich repeat‐containing receptors, whereas GPCR135 (RXFP3) and GPCR142 (RXFP4) resemble receptors that respond to small peptides such as somatostatin and angiotensin II. The cognate ligands for the receptors have been identified: relaxin for RXFP1; INSL3 for RXFP2; relaxin 3 for RXFP3 and INSL5 for RXFP4. RXFP1 and RXFP2 receptors produce increases in intracellular cAMP levels upon stimulation, although the response is complex and contains a component sensitive to PI‐3‐kinase inhibitors. There is also evidence that RXFP1 can activate Erk1/2 and nitric oxide synthase, and relaxin has been reported to enter cells and activate glucocorticoid receptors. In contrast, RXFP3 and RXFP4 couple to Gi by a pertussis toxin‐sensitive mechanism to cause inhibition of cAMP production. Now that the receptors for relaxin family peptides and their cognate ligands have been identified, we suggest a nomenclature for both the peptides and the receptors that we hope will be helpful to researchers in this rapidly advancing field.
Expression of Porcine Prorelaxin in Transgenic Tobacco
Authors: BUSWELL SCOTT , MEDINA‐BOLIVAR FABRICIO , CHEN QIANG , VAN COTT KEVIN , ZHANG CHENMING
Published: June 2005

Abstract: In vitro studies demonstrate that porcine relaxin may possess various therapeutic effects. In this study, we explore the possibility of expressing porcine relaxin in transgenic tobacco. Tobacco was selected because it is a non‐food, non‐feed crop, and recombinant protein production can readily be scaled up. The cDNA of porcine preprorelaxin was under the regulation of two different constitutive promoters. DNA analysis by polymerase chain reaction verified that all transgenic plants contained the correct size of gene insert. Preliminary studies showed the presence of putative prorelaxin bands in both silver‐stained SDS‐PAGE and western blot. The results also indicated that tobacco‐produced prorelaxin may not be properly processed to yield the mature relaxin.
Oxytocin and Estrogen Receptor Expression in the Myometrium of Pregnant Relaxin‐Deficient (Rlx−/−) Mice
Authors: SIEBEL ANDREW L. , GEHRING HELEN M. , VODSTRCIL LENKA , PARRY LAURA J.
Published: June 2005

Abstract: Relaxin decreases oxytocin‐stimulated rat myometrial contractions in vitro. This study used pregnant relaxin‐deficient (Rlx−/−) mice to investigate the interaction between relaxin, oxytocin receptor (OTR), and estrogen receptor (ER) expression in the myometrium. Myometrial OTRs were significantly decreased on gestation day 18.5 in Rlx−/− mice than in Rlx+/+ mice. An increase in ERα in Rlx+/+ mice at term was correlated with a decrease in ERβ, which was not observed in Rlx−/− mice. Treatment of Rlx−/− mice with relaxin had no effect on OTR, LGR7, or ERα expression, but it caused a significant decrease in ERβs.
Role of Relaxin in Maternal Renal Vasodilation of Pregnancy
Authors: CONRAD KIRK P. , JEYABALAN ARUN , DANIELSON LESLIE A. , KERCHNER LAURIE J. , NOVAK JACQUELINE
Published: June 2005

Abstract: The remarkable hemodynamic changes of normal pregnancy are briefly reviewed. In addition, new findings and current concepts related to the underlying hormonal and molecular mechanisms are presented. Finally, work that is in progress as well as future directions is briefly discussed.
Localization of LGR7 Gene Expression in Adult Mouse Brain Using LGR7 Knock‐out/LacZ Knock‐in Mice: Correlation with LGR7 mRNA Distribution
Authors: PICCENNA LORETTA , SHEN PEI‐JUAN , MA SHERIE , BURAZIN TANYA C. D. , GOSSEN JAN A. , MOSSELMAN SIETSE , BATHGATE ROSS A. D. , GUNDLACH ANDREW L.
Published: June 2005

Abstract: Knowledge of the distribution of the relaxin receptor, LGR7, in the brain provides a basis for studies of the physiologic actions of relaxin. LGR7 knock‐out (KO) mice were produced by the in‐frame replacement of LGR7 exon 10 and 11 with a LacZ‐reporter cassette (knock‐in [KI]), and in this study we used LGR7‐KO/LacZ‐KI mice to determine the regional/cellular distribution of LGR7 gene expression in adult mouse brain by assessing β‐galactosidase activity in perfusion‐fixed sections. High densities of β‐galactosidase‐positive neurons were detected in anterior olfactory and claustrum/endopiriform nuclei, deep layers of cortex (particularly somatosensory), and the subiculum. Low to moderate densities were detected in olfactory bulb (periglomerular layer), cingulate cortex, subfornical organ, hippocampal CA2/dentate hilus, amygdala, hypothalamus, and thalamus. This LGR7/LacZ expression appears to recapitulate that of native LGR7 in wild‐type mice and provides a model to further investigate the phenotype of LGR7‐responsive neurons in the brain and to help reveal functions associated with central relaxin signaling.
Relaxin Inhibits Central Angiotensin II Expression in Killifish: A Central Osmoregulatory Role for Relaxin and Angiotensin II in the Killifish Fundulus heteroclitus
Authors: AMAMOO ADWOA , WILSON BRIAN C.
Published: June 2005

Abstract: We investigated whether relaxin and brain angiotensin II (AII) are osmoregulatory hormones in killifish. Four groups of killifish acclimatized to different saline environments (freshwater, 25% sea water, 50% sea water [brackish], and sea water) were euthanized and their brains were embedded in paraffin. A separate group of brackish water killifish were injected with relaxin 1 h before dissection and embedding. Serial sagittal sections were immunohistochemically stained for AII. Angiotensin II immunostaining did not differ between saline environments. Contrary to expectation, relaxin decreased the number of AII‐immunoreactive cells in hypothalami of brackish water fish.
Relaxin Signaling from Natural Receptors
Authors: IVELL RICHARD , ANAND‐IVELL RAVINDER , BARTSCH OLAF
Published: June 2005

Abstract: The heterodimeric peptide hormone relaxin in most cells appears to signal through a G‐protein‐coupled receptor, LGR7. Whereas in artificial cell systems, made by transfection of receptor‐expressing gene constructs into cells normally not presenting the receptor, classic activation of adenylate cyclase appears to be mediated by Gs, in cells naturally expressing the receptor, this type of coupling appears to be very weak. Instead, there is good evidence of other intermediate steps involving cytoplasmic components and tyrosine kinase activity. Part of the complexity of relaxin signaling is also manifest in the variable time course of cAMP production evident in the THP‐1 cell line, which appears to depend on passage number and, hence, presumably on differentiation status. It is therefore important to distinguish between immediate early effects, short to mid‐term responses, and long‐term responses likely the consequences of specific gene upregulation.
Signal Switching after Stimulation of LGR7 Receptors by Human Relaxin 2
Authors: HALLS MICHELLE L. , BATHGATE ROSS A. , SUMMERS ROGER J.
Published: June 2005

Abstract: Previous studies have described a biphasic cAMP response after stimulation of LGR7 by human gene 2 (H2) relaxin, involving both adenylate cyclase and PI3‐kinase activity. The current study identifies the upstream involvement of Gi in the PI3‐kinase‐mediated response, likely the result of receptor signal switching. Amino acid sequence analysis of the LGR7 C‐terminal tail and intracellular loops revealed multiple putative phosphorylation sites, suggesting that signal switching from Gs to Gi may occur after receptor phosphorylation. This study supports a time‐dependent biphasic cAMP response: an initial short Gs‐adenylate cyclase‐mediated cAMP response is followed by receptor signal switching to a Gi‐PI3‐kinase‐mediated response.
Relaxin Restores Altered Ileal Spontaneous Contractions in Dystrophic (mdx) Mice
Authors: BACCARI MARIA CATERINA , CALAMAI FRANCO , CHIAPPINI LAURA , VANNUCCHI MARIA GIULIANA , BANI DANIELE
Published: June 2005

Abstract: We studied the effects of relaxin on ileal contractility in normal and dystrophic (mdx) mice. Ileal preparations from male normal and mdx mice showed spontaneous myogenic contractions whose amplitude was significantly higher in the latter ones. Relaxin added to the bath medium together with l‐arginine depressed the amplitude of the spontaneous contractions in the mdx mice to a level similar to that of the normal mice. The nitric oxide synthase (NOS) inhibitor L‐NG‐nitroarginine reverted this effect. In mdx mice pretreated for 18 hours with relaxin, spontaneous motility was greatly reduced in amplitude, resembling that of the normal mice. Concurrently, iNOS expression in the muscle coat was markedly increased. Therefore, in mdx mice, relaxin can restore an ileal motility pattern similar to that of the normal mice by upregulating endogenous NO biosynthesis.
Relaxin Induces Matrix Metalloproteinase‐9 through Activation of Nuclear Factor Kappa B in Human THP‐1 Cells
Authors: HO TEH‐YUAN , BAGNELL CAROL A.
Published: June 2005

Abstract: Matrix metalloproteinase (MMP) and relaxin are important for tissue remodeling and wound repair. Macrophages populate wound sites and secrete MMPs. Nuclear factor kappa B (NF‐κB) is linked to MMP gene regulation. Thus, a monocyte/macrophage cell line, THP‐1, was used to study the mechanism of relaxin's action on MMPs. Relaxin increased MMP‐9 protein and activity in THP‐1 cell‐conditioned media, with no significant change in MMP‐2 activity. NF‐κB DNA binding activity was elevated in response to relaxin, and supershift assay showed activation of both NF‐κB subunits p50 and p65. Relaxin also reduced NF‐κB inhibitor protein, IκB‐α. In conclusion, these data suggest that relaxin‐induced MMP‐9 expression in THP‐1 cells involves NF‐κB activation.
Does Human Relaxin‐2 Affect Peripheral Blood Mononuclear Cells to Increase Inflammatory Mediators in Pathologic Bone Loss?
Authors: KRISTIANSSON P , HOLDING C , HUGHES S , HAYNES D
Published: June 2005

Abstract: This study was designed to test the hypothesis that relaxin stimulates bone resorption by regulating the production of several mediators that stimulate osteoclast formation. The levels of mediators were measured in response to differing relaxin concentrations in supernatants from peripheral blood mononuclear cells (PBMCs), MCF‐7 breast cancer cells, and normal human osteoblasts. Although all cell types expressed mRNA for the relaxin receptor (LGR7), only PBMCs responded to relaxin at physiologic levels by increasing tumor necrosis factor‐α and interleukin‐1β secretion. The findings indicate that PBMCs should be studied in relation to the effect of relaxin on inflammation and bone destruction caused by osteoclasts.
Demonstration of Upregulated H2 Relaxin mRNA Expression during Neuroendocrine Differentiation of LNCaP Prostate Cancer Cells and Production of Biologically Active Mammalian Recombinant 6 Histidine‐Tagged H2 Relaxin
Authors: FIGUEIREDO KEVIN A. , PALMER JODIE B. , MUI ALICE L. , NELSON COLLEEN C. , COX MICHAEL E.
Published: June 2005

Abstract: Relaxin was recently implicated as a regulator of breast and prostate cancer progression. We characterized upregulated H2 relaxin gene expression during neuroendocrine differentiation of the human prostate cancer model, LNCaP. To examine the impact of relaxin on host cells associated with prostatic adenocarcinomas, we generated recombinant 6 His‐tagged relaxin (RLXH) in a mammalian expression system. This immunoreactive and biologically active relaxin preparation was used to screen a variety of cell types for cAMP responsiveness. Of the cell types screened, none was more responsive to RLXH than the well‐characterized monocyte/macrophage cell line THP‐1.
Responses of GPCR135 to Human Gene 3 (H3) Relaxin in CHO‐K1 Cells Determined by Microphysiometry
Authors: VAN DER WESTHUIZEN EMMA T. , SEXTON PATRICK M. , BATHGATE ROSS A. D. , SUMMERS ROGER J.
Published: June 2005

Abstract: This study examined the functional response to human relaxin 2 (H2 relaxin), human relaxin 3 (H3 relaxin), porcine relaxin, and human INSL3 in the cytosensor microphysiometer, using CHO‐K1 cells stably expressing human GPCR135. CHO‐K1 cells stably expressing GPCR135 were generated by the serial dilution method and receptor properties were assessed. Saturation studies of [125I] H3 relaxin binding to GPCR135 in these cells gave a Bmax of 32.61 ± 6.5 fmol/mg protein and Kd of 0.12 ± 0.08 nM. The functional response to H3 relaxin and other relaxin/insulin peptides of GPCR135 expressed in CHO‐K1 cells was measured in the cytosensor microphysiometer and analyzed using inhibitors of signal transduction proteins.
Mechanisms of Relaxin‐Mediated Premature Birth
Authors: WEISS GERSON , GOLDSMITH LAURA T.
Published: June 2005

Abstract: In women, circulating relaxin is produced by the corpus luteum of pregnancy. The levels of relaxin are predominantly determined by the luteal mass, the number of corpora lutea present. Relaxin levels are highest after ovulation induction, which stimulates formation of many corpora lutea. Elevated relaxin levels in the first trimester of pregnancy are maintained throughout pregnancy and are linearly related to preterm birth. In an in vitro model of late human pregnancy cervix, relaxin increases MMP‐1 and MMP‐3 and decreases TIMP‐1 levels, thus acting as a positive regulator of matrix metalloproteinases. In an in vivo rhesus monkey model of early pregnancy, relaxin decreases cervical collagen content, decreases cervical lumican levels, and stimulates MMP‐7 levels. Early effects of relaxin in the uterus include increasing endometrial arteriole number and increasing the number of leukocytes, uterine natural killer cells, macrophages, and neutrophils. These cells release many cytokines which contribute to changes that stimulate and facilitate uterine contractility. If these changes persist in late pregnancy, relaxin may be a mediator of labor. Excess relaxin may produce these changes at an accelerated rate, causing preterm birth.
Use of Relaxin in Orthodontics
Authors: STEWART DENNIS R. , SHERICK PAUL , KRAMER SUSAN , BREINING PETER
Published: June 2005

Abstract: BAS Medical is investigating the use of relaxin to improve ortho‐dontic treatments. Relaxin is well known for its effects on remodeling soft tissue, and we believe relaxin will be clinically useful in speeding tooth movement and preventing relapse. We investigated the use of relaxin in preventing relapse in a dog model. Dog maxillary second incisors were orthodontically rotated an average of 42 degrees, and then relaxin was administered by gingival injection to relieve the rotational memory in the connective tissues. Teeth were retained for 30 days to allow fibers to reform. Teeth then were released and relapse was measured by a series of impressions. Animals receiving relaxin gingival injections (n= 8) were compared with placebo‐treated animals (n= 8) (exhibiting high relapse) and gingival fiberotomies (n= 8) (low relapse). Gingival fiberotomy is a surgical procedure to cut the gingival connective tissues away from the tooth and has been shown to be effective in preventing relapse clinically and in animal models. There was a significant difference in relapse between the fiberotomy and the placebo control groups, and the relaxin‐treated group had an intermediate response between the two groups (nonsignificant). Dose and treatment optimization may improve the response in future studies. To study the underlying mechanisms, we have localized relaxin receptors on gingival and periodontal ligament fibroblasts in tissue slices and cell cultures. Relaxin was found to stimulate collagenase production by relaxin in human gingival fibroblast cultures. Taken together, the data support a role for relaxin therapy to speed tooth movement and prevent relapse in orthodontic practice.
The Use of Relaxin Improves Healing in Injured Muscle
Authors: LI Y , NEGISHI S , SAKAMOTO M , USAS A , HUARD J
Published: June 2005

Abstract: To improve the functional recovery of injured skeletal muscle, we have focused our efforts on both enhancement of muscle regeneration and prevention of fibrosis. The polypeptide cytokine/growth factor relaxin can inhibit fibrous tissue formation in many tissues. As a member of the insulin‐like growth factor family, relaxin also is a potential stimulator of muscle regeneration. In the current experiment, we examined the antifibrotic effect of relaxin in injured skeletal muscle. We also investigated if the injection of relaxin would influence muscle regeneration after injury. Our results demonstrate that relaxin treatment improved histologic and physiologic healing of muscles subjected to traumatic injury.
Human Medullary Thyroid Carcinoma: A Source and Potential Target for Relaxin‐Like Hormones
Authors: KLONISCH T , MUSTAFA TAREK , BIALEK JOANNA , RADESTOCK YVONNE , HOLZHAUSEN HANS‐JÜRGEN , DRALLE HENNING , HOANG‐VU CUONG , HOMBACH‐KLONISCH SABINE
Published: June 2005

Abstract: We investigated the expression of H1, H2 relaxin and INSL‐3, mRNA and protein, and LGR7 and LGR8 transcripts in human C‐cell hyperplasia, primary medullary thyroid carcinoma (MTC) tissues, MTC metastases, and the human MTC‐TT and mouse MTC‐M cell lines. Relaxin‐like peptide hormones were detected in C‐cell hyperplasia and in MTC tissues, but were absent in human normal parafollicular C‐cells of benign goiter tissues. In contrast to calcitonin, mRNA, and immunoreactive protein, no differences in the expression of relaxin and INSL3 were observed in MTC tissues of different pTNM classification or between primary and metastatic MTC tissues studied. All MTC tissues constitutively expressed LGR7 and LGR8 transcripts. Thus, relaxin‐like hormones appear to be present early during C‐cell hyperplasia and potentially functional relaxin/INSL3 ligand‐receptor systems are present in human MTC tissues and in MTC cell lines.
Relaxin Downregulates the Calcium Binding Protein S100A4 in MDA‐MB‐231 Human Breast Cancer Cells
Authors: RADESTOCK YVONNE , HOANG‐VU CUONG , HOMBACH‐KLONISCH SABINE
Published: June 2005

Abstract: Expressed in the human breast and in human breast cancer tissues, the heterodimeric peptide hormone relaxin is involved in extracellular matrix turnover. To investigate the role of relaxin in estrogen receptor‐alpha negative human breast cancer cells, we established transfectants of the human MDA‐MB‐231 breast cancer cell line stably overexpressing H2‐relaxin (MDA‐MB‐231/pIRES‐EGFP‐H2). These transfectants produced and secreted functional relaxin. Our investigations revealed downregulation of mRNA and protein of the calcium binding protein S100A4 (metastasin) in MDA‐MB‐231/pIRES‐EGFP‐H2 transfectants, indicating a distinct and novel role for relaxin in estrogen receptor‐alpha negative human breast cancer cells.
Effects of Relaxin in a Model of Rat Adjuvant‐Induced Arthritis
Authors: SANTORA KAREN , RASA CORDELIA , VISCO DENISE , STEINETZ BERNARD , BAGNELL CAROL
Published: June 2005

Abstract: A reduction in the incidence and severity of rheumatoid arthritis is seen in pregnant women. Relaxin, a hormone of pregnancy, has been implicated in decreased immune responsiveness. Consequently, the effects of relaxin and estradiol valerate, alone or in combination, were assessed in the development of adjuvant‐induced arthritis in the rat. Combination hormone therapy reduced adjuvant‐induced paw inflammation. Radiographic analysis of the tarsal joints showed that estradiol valerate plus relaxin treatment minimized soft tissue damage and bone changes when compared to vehicle‐treated arthritic controls. These results indicate that relaxin may be a factor in reducing inflammation during pregnancy.