Aspectos da biologia reprodutiva de Boa constrictor constrictor: um estudo histológico dos testículos nos períodos reprodutivos de quiescência e máxima atividade / Boa constrictor constrictor reproductive biology aspects: the testes histological study in the reproductive periods of quiescence and maximal activity

As serpentes pertencem ao segundo maior grupo dentro dos répteis, podendo apresentar sazonalidade quanto à espermatogênese, com produção descontínua ou contínua. O presente trabalho tem como objetivo caracterizar aspectos da biologia reprodutiva de Boa constrictor constrictor com base nos achados histológicos dos testículos nos períodos de máxima atividade (período de gametogênese) e quiescência. Os testículos de dois espécimes de Boa c. constrictor (7767 e 11752) foram cortados a uma espessura de 3µm em micrótomo, corados com azul de toluidina 1%, fotodocumentados e descritos. A presença de espermatozoides na luz do túbulo seminífero no indivíduo 7767 indica um período de máxima gametogênese, enquanto o lúmen dos túbulos seminíferos pouco evidentes, sem a presença de espermatozoides e de células gaméticas em divisão, caracteriza o indivíduo 11752 em período quiescente. Mediante os achados histológicos descritos no presente estudo, concluiu-se que Boa c. constrictor apresenta sazonalidade em relação à gametogênese, sendo esse padrão de sazonalidade associado ao período de cópulas relatado em literatura característico de serpentes com padrão pré-nupcial.(AU)

These snakes belong to the second largest group within the reptiles, being able to present seasonality regarding spermatogenesis, with discontinuous or continuous production. The present study aims to characterize Boa constrictor constrictor reproductive biology aspects from histological findings in testicles during periods of maximum activity (period of gametogenesis) and quiescence. The testicles of two specimens of Boa c. constrictor (7767 and 11752) were cut to a thickness of 3µm in microtome, stained with 1% toluidine blue, photodocumented and described. The spermatozoa presence in the seminiferous tubule lumen in individual 7767 indicates a period of maximum gametogenesis, whereas the seminiferous tubules lumen is not very evident without spermatozoa and the absence of dividing gametic cells characterizes individual 11752 in the quiescent period. Through the histological findings we concluded that Boa c. constrictor presents seasonality in relation to gametogenesis, and the pattern of reproductive seasonality observed along with the period of copulas reported in the literature resembles the pre-nuptial pattern.(AU)

Anti-ß2-glycoprotein I and anti-phosphatidylserine/prothrombin antibodies exert similar pro-thrombotic effects in peripheral blood monocytes and endothelial cells.

PURPOSE: The introduction of the anti-phosphatidylserine/prothrombin (aPS/PT) antibodies among the routinely investigated anti-phospholipid (aPL) antibodies led to an improvement in anti-phospholipid syndrome (APS) laboratory diagnostic performance; however, their pathogenic mechanism is still substantially undefined. To support clinical data and future inclusion as possible new criteria antibodies, we designed a head-to-head study to directly compare the procoagulant effects sustained in vitro by aPS/PT to those sustained by anti-ß2-glycoprotein I (aß2GpI) domain 1-specific antibodies. METHODS: Blood donors-derived monocytes and endothelial cells (HUVEC) were stimulated with lipopolysaccharides (LPS) alone or in combination with the IgG fractions isolated from the serum of six APS patients, positive only for aß2GpI or for aPS/PT antibodies. As control, cells were incubated with LPS plus the IgG isolated from blood donors. Tissue factor (TF) mRNA expression was measured after four hours incubation by real-time PCR. Nitric oxide (NO) levels were measured in cells supernatant after 16 h incubation by colorimetric assay. RESULTS: aPS/PT and aß2GpI IgG antibodies fractions showed comparable ability to enhance LPS-induced TF mRNA expression, either in monocytes and in HUVEC. Compared to LPS alone, we found that NO levels are strongly overproduced in HUVEC treated with LPS plus aß2GpI and aPS/PT IgG fractions. CONCLUSIONS: Our data support the significant and independent role of aPS/PT in the pathogenesis of the thrombotic events in APS patients, possibly adding new light to the therapeutic management of cases characterized by the sole presence of aPS/PT IgG antibodies.

A versatile method for gene dosage quantification: multiplex PCR and single base extension for copy number and gene-conversion identification of SMN genes.

A comparison of the individual genomes within a species demonstrates that structural variation, including copy number variation (CNV), is a major contributor to phenotypic diversity and evolutionary adaptation. CNVs lead to the under/over-expression of a gene, according to the changes in the gene dosage, which account for the development of a number of genomic disorders. Thus, the development of efficient, rapid and accurate CNV screening is of fundamental importance. We report a method that enables the simultaneous determination of the copy numbers of several different targets as well as the discrimination among highly similar/almost identical targets that differ by only one single nucleotide variant, which establishes their copy numbers. The PCR co-amplification and single-base extension technologies are used to identify the copy number of a target sequence relative to a reference sequence of known genomic copy number in a given sample. This efficient and accurate quantification platform was successfully used to quantify the copy numbers of the primary spinal muscular atrophy-determining gene, SMN1, and the disease modifier gene, SMN2. The reliability, low-cost and potential for high-throughput make our method suitable for screening large populations as well as for use as a tool in clinical settings for genetic diagnosis/prognosis.

The -308 TNFα and the -174 IL-6 promoter polymorphisms associate with effective anti-TNFα treatment in seronegative spondyloarthritis.

The genetic predisposition to a long-term efficacy of anti-tumor necrosis factor (TNF)α treatment in seronegative spondyloarthritis (SpA) was investigated by analysing the possible correlation between several single nucleotide gene polymorphisms and the retention rate of anti-TNFα therapies. We compared patients needing to switch the first anti-TNFα (Sw, No. 64) within at least 12 months of follow-up with patients not needing to switch (NSw, No. 123), observing at least 6 months of treatment to establish anti-TNFα failure, leading to treatment change. Response to treatment was evaluated by standardised criteria (BASDAI for axial involvement, DAS28-EULAR for peripheral involvement). The TNFα -308 A allele and the interleukin (IL)-6 -174GG homozygosis resulted as independent biomarkers predicting survival of the first anti-TNFα therapy in SpA patients (P=0.007, odds ratio (OR): 4.4, 95% confidence interval (CI)=1.5-13.1 and P=0.035, OR: 2.1, 95% CI=1.1-4.4). Also, the male gender (P=0.001, OR: 3.4, 95% CI=1.6-7.1) associated with the NSw phenotype, whereas no association was found either with the specific diagnosis or the predominant joint involvement.

Second trimester amniotic fluid retinol in patients developing preeclampsia.

BACKGROUND: Retinol (ROH) is an essential micronutrient required for normal fetal development and an essential molecule for antioxidant processes. OBJECTIVE: To investigate the putative role of ROH as a marker of preeclampsia in early second trimester amniotic fluid (AF). MATERIALS AND METHODS: Case-control study comparing the concentration of ROH and other antioxidants such as uric acid, vitamin E and malondialdehyde (MDA) in second trimester AF in patients that later developed preeclampsia with normal pregnancies. RESULTS: The concentration of ROH in amniotic fluids of women that later developed preeclampsia was significantly higher than those of uncomplicated pregnancies (66.72 µg/l (49.00-70.56) vs. 44.4 µg/l (31.9-51.17), p < 0.05). No statistical significant difference was found in uric acid, vitamin E and MDA concentration. In the multivariate logistic regression, concentrations of ROH in amniotic fluids directly correlate with the risk of developing preeclampsia (OR 1.13, IC 0.01-1.26, p < 0.05). CONCLUSIONS: Second trimester AF ROH concentration was significantly higher in pregnancies that developed preeclampsia compared to normal pregnancies.

Reinterpretation of mouse thyroid changes under space conditions: the contribution of confinement to damage.

During space missions, astronauts work in a state of separation from their daily social environment and in physical confinement. It has been shown that confinement influences mood and brain cortical activity, but no data has been obtained with regard to its effect on the thyroid gland, the structure and function of which change during spaceflights. Here, we report the results of a study on the effects of confinement on mouse thyroid, which was implemented with the Mice Drawer System Facility maintained on the ground, a system used for spaceflight experiments. The results show that confinement changes the microscopic structure of the thyroid gland and that it exhibits symptoms similar to those that result from physiological and/or pathological hyperfunction. What is left unchanged, however, is the sphingomyelinase-thyrotropin receptor relationship, which is important for thyrotropin response with a consequential production of hormones that act on the metabolism of almost all tissues and reduces the production of calcitonin, a hormone involved in bone metabolism. During space missions, the overexpression of pleiotrophin, a widespread cytokine up-regulated after tissue injury that acts on bone remodeling, attenuates changes to the thyroid that are spaceflight-dependent; therefore we studied the thyroids of pleiotrophin-transgenic mice in the Mice Drawer System Facility. In confinement, pleiotrophin overexpression does not protect from the loss of calcitonin. The contribution of confinement to thyroid damage during spaceflights is discussed.

Observing the mouse thyroid sphingomyelin under space conditions: a case study from the MDS mission in comparison with hypergravity conditions.

This is a case report of apparent thyroid structural and functional alteration in a single mouse subjected to low Earth orbit spaceflight for 91 days. Histological examination of the thyroid gland revealed an increase in the average follicle size compared to that of three control animals and three animals exposed to hypergravity (2g) conditions. Immunoblotting analysis detected an increase in two thyroid gland enzymes, sphingomyelinase and sphingomyelin-synthase1. In addition, sphingomyelinase, an enzyme confined to the cell nucleus in the control animals, was found in the mouse exposed to hypogravity to be homogeneously distributed throughout the cell bodies. It represents the first animal observation of the influence of weightlessness on sphingomyelin metabolism.

The thyroid lobes: the different twins.

Although differences in size of the right and left thyroid lobes are well defined, differences in morphology, follicles structure, cAMP production, thyrotropin receptor, and protein involved in cell signalling have not previously been reported. This study provides morpho-functional data of right and left thyroid lobes by biochemical, immunohistochemistry, immunoblotting and immunofluorescence analysis. We demonstrate that, in comparison with the left lobe, the right lobe has a higher activation index, is more sensitive to thyrotropin treatment, is rich in thyrotropin receptor and caveolin 1 involved in thyroid hormone synthesis as well as in epithelial thyroid cell homeostasis, is characterised by a high content of molecules involved in cell signalling such as stat3, raf1, sphingomyelinase and sphingomyelin-synthase whose activity ratio is necessary for epithelial cell activity and finally has more areas calcitonin-dependent. The relation between structure/function of right lobe and its susceptibility to the higher risk of pathological modifications with respect the left lobe is discussed.

Buprenorphine/naloxone versus methadone in opioid dependence: a longitudinal survey.

BACKGROUND AND OBJECTIVES: Buprenorphine and methadone are widely used for the treatment of opioid dependence, but their diversion and/or misuse are frequent. In principle, buprenorphine/naloxone combination therapy should be associated with a lower frequency of drug abuse/misuse than methadone. This study assessed the efficacy of the substitution of buprenorphine treatment with the buprenorphine/naloxone combination in opioid-dependent patients. MATERIAL AND METHODS: 3812 drug-addicted outpatients selected from 10 Italian Public Services for Addiction (Ser.T.) centres in Naples (Italy) were enrolled: 3105 (81.5%) were treated with methadone and 707 (18.5%) with buprenorphine. The buprenorphine treatment was switched to buprenorphine/naloxone (4:1), and the patients were followed for about 1 year. The number of subjects still on treatment after 1 year, their status according to social, educational and toxicologic (assessed by a urine toxicology test) parameters were assessed. RESULTS: 1 year after the therapy switch, the number of patients still on treatment was similarly reduced with methadone (2883; -7.5%) and buprenorphine/naloxone (632; -10.6%; p=0.369). However, in patients treated with buprenorphine/naloxone, a significant improvement was reported in social life status (63% versus 39% of the buprenorphine/naloxone and methadone treated subjects, respectively, were married/cohabiting p<0.001), in the educational level (43% of buprenorphine/naloxone treated versus 32% of the methadone treated subjects obtained at least a high school certificate, p<0.001) and in the toxicological conditions (53% of buprenorphine/naloxone treated subject versus 30% of methadone treated individuals had opioid- and cocaine- negative urine tests, p<0.001). DISCUSSION: These preliminary data suggest that buprenorphine/naloxone treatment of opioid dependence reduces the percentage of treated subjects similarly to methadone, and is associated with an improvement in social life, educational and toxicological conditions, compared with methadone treatment. However, we cannot exclude a selection bias, i.e. patients who were more likely to stabilize their opiate dependence switched to buprenorphine/naloxone.

Thyrotropin receptor and membrane interactions in FRTL-5 thyroid cell strain in microgravity.

The aim of this work was to analyze the possible alteration of thyrotropin (TSH) receptors in microgravity, which could explain the absence of thyroid cell proliferation in the space environment. Several forms of the TSH receptor are localized on the plasma membrane associated with caveolae and lipid rafts. The TSH regulates the fluidity of the cell membrane and the presence of its receptors in microdomains that are rich in sphingomyelin and cholesterol. TSH also stimulates cyclic adenosine monophosphate (cAMP) accumulation and cell proliferation. Reported here are the results of an experiment in which the FRTL-5 thyroid cell line was exposed to microgravity during the Texus-44 mission (launched February 7, 2008, from Kiruna, Sweden). When the parabolic flight brought the sounding rocket to an altitude of 264 km, the culture media were injected with or without TSH in the different samples, and weightlessness prevailed on board for 6 minutes and 19 seconds. Control experiments were performed, in parallel, in an onboard 1g centrifuge and on the ground in Kiruna laboratory. Cell morphology and function were analyzed. Results show that in microgravity conditions the cells do not respond to TSH treatment and present an irregular shape with condensed chromatin, a modification of the cell membrane with shedding of the TSH receptor in the culture medium, and an increase of sphingomyelin-synthase and Bax proteins. It is possible that real microgravity induces a rearrangement of specific sections of the cell membrane, which act as platforms for molecular receptors, thus influencing thyroid cell function in astronauts during space missions.

UVC radiation-induced effect on human primary thyroid cell proliferation and HLA-DR expression.

The aim of this study was to examine how UVC irradiation will affect normal human thyroid cell proliferation and HLA-DR expression. Primary human thyroid cells were exposed to UVC (254 nm wavelength) irradiation. In some experiments 0.5 mM buthionine sulfoximine (BSO) was added. Apoptosis was detected measuring annexin V, proteins involved in apoptotic process (p53, Bax, Bcl-2, caspase 3, and 9) by immunoblot analysis and HLA-DR expression by FACS. UVC induced a cell cycle arrest in G0/G1 phase in the first 24 h, accumulation of cells in the S phase 72 h after treatment, and an increase of apoptotic cells. BSO pretreatment showed an earlier appearance and a higher percentage of apoptosis. p53, caspase 3 and 9 were increased, while Bax and Bcl-2 were decreased. We also observed a transient significant increase in HLA-DR expression. UVC inhibited cell proliferation and induced apoptosis in normal human primary thyroid cells. An inhibitor of glutathione synthesis induced an earlier appearance and higher percentage of apoptosis suggesting that oxidative stress may play a role. Apoptotis involved components of the intrinsic mitochondrial pathway. A transient increase in HLA-DR expression after UVC irradiation could play a role in inducing AITD.

Study on the possible role of the -174G>C IL-6 promoter polymorphism in predicting response to rituximab in rheumatoid arthritis.

OBJECTIVE: Identification of genetic biomarkers of response to biologics in rheumatoid arthritis (RA) is a relevant issue. The -174G>C interleukin-6 (IL-6) promoter polymorphism was investigated in RA patients treated with rituximab (RTX), being IL-6 a key cytokine for B cell survival and proliferation, thus possibly implicated in rituximab efficacy. METHODS: The study was conducted in a real-life retrospective cohort of 142 unselected RA patients (120F/22M) treated with RTX and referred to 7 rheumatologic centres in the north of Italy. One hundred and thirteen (79.6%) patients were rheumatoid factor (RF)-positive and 112 (78.9%) were anti-CCP antibodies positive. The response to therapy was evaluated at the end of the sixth month after the first RTX infusion, by using both the EULAR criteria (DAS28) and the ACR criteria. The IL-6 -174G>C promoter polymorphism was analyzed by RFLP following previously reported methods. RESULTS: Lack of response to RTX at month +6 by EULAR criteria was more prevalent in RA patients with the IL-6 -174 CC genotypes (9/21, 42.8%), than in the GC/GG patients (23/121, 19.0%) (OR 3.196, 95% CI=1.204-8.485; p=0.0234). Similar results were found when evaluating the response by ACR criteria. No differences were found in RA duration, baseline DAS28, baseline HAQ, RF status, anti-CCP status according to the different IL-6 -174 genotypes. CONCLUSION: IL-6 promoter genotyping may be useful to better plan treatment with RTX in RA. Larger replication studies are in course to confirm these preliminary results.

[Multicenter study on the monitoring of in vitro susceptibility to tigeeyeline in Santiago, Chile]. / Estudio multicéntrico de la vigilancia de la susceptibilidad in vitro a tigeciclina en Santiago de Chile.

The objective of this multicenter study was to determine tigecycline susceptibility rates, measured by agar diffusion, in nine hospitals in Santiago and to compare these rates with other antimicrobials. Each center studied 20 strains per month. All intermediate and fully resistant strains as well as 10% of susceptibile strains were also studied by the broth microdilution method. Overall, 2301 strains were studied displaying the following susceptibility rates for tigecycline: 100% for Streptococcus sp, Enterococcus sp, and E. coli respectively, 99.8% for Staphylococcus sp, 93% for Klebsiella and 80% for Acinetobacter baumarmii. For Proteus, Providencia and Morganella the susceptibility rates were 4%. For cefotaxime-resistant Klebsiella and imipenem-resistant A. baumarmii susceptibility rates were 95% and 80% respectively. The agar diffusion and broth dilution method were 100% concordant for tigecycline susceptible strains but only 27% for resistant or intermediate strains represented mostly by Acinetobacter baumannii. The majority of these strains (57/59) proved to be susceptible after retesting. The great majority (96,6%) of strains tested from nine Chilean hospitals proved to be susceptible to tigecycline with exception for Proteus, Providencia and Morganella (66% resistance). Using the agar diffusion method for measuring tigecycline susceptibility to A. baumannii may be misleading.

Estudio multicéntrico de la vigilancia de la susceptibilidad in vitro a tigeciclina en Santiago de Chile / Multicenter study on the monitoring of in vitro susceptibility to tigeeyeline in Santiago, Chile

The objective of this multicenter study was to determine tigeeyeline susceptibility rates, measured by agar diffusion, in nine hospitals in Santiago and to compare these rates with other antimicrobials. Each center studied 20 strains per month. All intermedíate and fully resistant strains as well as 10 percent of susceptibile strains were also studied by the broth microdilution method. Overall, 2301 strains were studied displaying the foliowing susceptibility rates for tigeeyeline: 100 percent for Streptococcus sp, Enterococcus sp, and E. coli respectively, 99.8 percent for Staphylococcus sp, 93 percent for Klebsiella and 80 percent for Acinetobacter baumarmii. For Proteus, Providencia and Morganella the susceptibility rates were 4 percent. For cefotaxime-resistant Klebsiella and imipenem-resistant A. baumarmii susceptibility rates were 95 percent and 80 percent respectively. The agar diffusion and broth dilution method were 100 percent concordant for tigeeyeline susceptible strains but only 27 percent for resistant or intermedíate strains represented mostly by Acinetobacter baumannii. The majority of these strains (57/59) proved to be susceptible after retesting. The great majority (96,6 percent) of strains tested from nine Chilean hospitals proved to be susceptible to tigeeyeline with exception for Proteus, Providencia and Morganella (66 percent resistance). Using the agar diffusion method for measuring tigeeyeline susceptibility to A. baumannii may be misleading.

Para conocer la susceptibilidad a tigeciclina por difusión en agar en nueve hospitales de Santiago y comparar la susceptibilidad con otros antimicrobianos, se diseñó este estudio multicéntrico. Cada centro estudió 20 cepas mensualmente. Las intermedias, resistentes y 10 por cientoo de las susceptibles se re-testearon y estudiaron por microdilución en caldo. Se incluyeron 2.304 cepas. Fueron susceptibles a tigeciclina Strep-tococcus sp (100 por cientoo), Enterococcus sp (100 por ciento), E. coli (100 por cientoo), Staphylococcus sp (99,8 por ciento), Klebsiella pneumoniae (93 por ciento) y Acinetobacter baumannii (80 por ciento). En Proteus, Providencia y Morganella la susceptibilidad fue 4 por cientoo. Klebsiella resistente a cefotaxima y Acinetobacter resistente a imipenem, 95 por cientoo y 80 por cientoo fueron susceptibles a tigeciclina, respectivamente. La concordancia en cepas susceptibles y en las enviadas como resistentes o intermedias (A. baumannii) fue 100 por cientoo y 27 por cientoo respectivamente. El re-testeo confirmó que la mayoría eran susceptibles. Los patrones de susceptibilidad bacteriana muestran muy buena actividad in vitro a tigeciclina. La resistencia in vitro de A. baumannii por difusión en agar debe interpretarse con precaución.

Differential expression of the components of the plasminogen activating system in human thyroid tumour derived cell lines and papillary carcinomas.

We characterised the expression of the plasminogen activators (uPA and tPA), the uPA receptor (uPAR) and the PAs inhibitors (PAI-1 and PAI-2) in human thyroid cell lines derived from normal thyroid, follicular adenoma, follicular, papillary and anaplastic carcinomas. Urokinase PA activity was detected in the supernatant of normal thyrocytes and augmented in those of all tumour cells. Quantitative RT-PCR analysis showed that uPA, uPAR and PAI-1 mRNAs increased in all carcinoma cells. Similar results were found in 13 papillary thyroid carcinoma (PTC) tissues which were mirrored in Western blot experiments. A correlation was found between tumour size and uPA mRNA increase, and higher levels of uPA and uPAR mRNAs were found in metastatic PTC. In conclusion, thyroid carcinoma cell lines and PTC overexpress uPA, uPAR and PAI-1 and the correlation of uPA and its cognate receptor with tumour size and metastasis may suggest their potential prognostic relevance in thyroid cancer.

Expression of matrix metalloproteinases and their specific inhibitors in normal and different human thyroid tumor cell lines.

In the present study we investigated, by means of zymography and reverse transcription-polymerase chain reaction (RT-PCR), the expression of different matrix metalloproteinases (MMPs) and of the specific tissue inhibitor of metalloproteinases [TIMPs] in human cell lines derived from normal thyrocytes (HTU5), follicular adenoma (HTU42), and follicular (FTC-133), papillary (B-CPAP), and anaplastic (CAL-62, 8305C) thyroid carcinomas. We demonstrated that normal thyrocytes constitutively express MMP-1, MMP-2, MMP-10, MMP-14, and TIMP-1, TIMP-2, TIMP-3, and TIMP-4, and this pattern of expression is profoundly modified in all thyroid tumor-derived cell lines. Analysis of the gelatinolytic activity in the different cell supernatants showed that the expressions of MMP-2 and MMP-9 are, respectively, increased or induced in all the neoplastic cell lines, except in CAL-62. Caseinolytic activity was found only in the supernatants of the 8305C and B-CPAP cells. Using RTPCR analysis we detected an increased expression of MMP-1 in cell lines derived from papillary and from one (8305C) of the two anaplastic carcinomas. MMP-13 mRNA was expressed only in the 8305C, FTC-133, and BCPAP cells. Among stromelysins, MMP-3 mRNA could not be detected in any cell line, while MMP-10 mRNA was expressed in all of them, although at variable levels. MMP-11 mRNA was absent in normal and follicular adenoma derived thyrocytes and induced in all carcinoma cell types. The expression of MMP-14 (MT1-MMP) mRNA was found significantly increased in all thyroid tumor cell lines with respect to HTU5 and HTU42 cells. The expression of TIMP-1 and TIMP-2 mRNAs was maintained in all cell lines tested, while that of TIMP-3 was lost in both anaplastic carcinoma cell lines and that of TIMP-4 was absent in the CAL-62. In conclusion, our data demonstrated a differential expression of MMPs and TIMPs in different thyroid tumor cell types with respect to normal thyrocytes. In particular, the induction of MMP-11 in all thyroid-derived carcinoma cell lines studied and of MMP-13 in all but one may represent, if confirmed in other thyroid tumor-derived cell lines and in thyroid tumor tissues, a new marker of thyrocyte transformation.

Atrial natriuretic factor and C-type natriuretic peptide induce retraction of human thyrocytes in monolayer culture via guanylyl cyclase receptors.

The natriuretic peptides signal through three receptor subtypes, of which two (NPR-A and NPR-B) are membrane-bound guanylyl cyclases for which the principal ligands are respectively atrial natriuretic factor (ANF) and C-type natriuretic peptide (CNP). In the human thyroid cell, a third receptor, NPR-C, has been implicated in the regulation of thyroglobulin, but functional roles for NPR-A and NPR-B have not yet been defined. In the present study we used RT-PCR to identify transcripts of all three receptor subtypes, both in human thyroid and in HTU-5 cells, a long-term culture of thyroid-derived cells. Both ANF and CNP induced a twofold increase in intracellular cGMP content in HTU-5 cells. Morphologic changes (a significant increase in cells of the retracted phenotype) were observed in ANF- and CNP-treated cells within 3 and 5 h of treatment respectively. Significant increases in retracted cell number were induced by ANF and CNP, but not the NPR-C-specific ring-deleted ANF analog, C-ANF(4-23), during a 15-day treatment. All three natriuretic peptides, however, induced a small (15-20%) but significant (P<0 small middle dot001) increase in DNA content per well. The stable analog of cGMP, 8-bromo-cGMP (8-BrcGMP; 1 mM), also increased the number of retracted HTU-5 cells, and was equipotent with the cAMP analog, 8-BrcAMP, in this effect. The cGMP-dependent protein kinase inhibitor, KT5823, however, had no significant effect on the ANF-induced increase in numbers of retracted cells. These results suggest that the actions of NPR-A and NPR-B, functional receptors in the human thyroid cell, may in part be mediated by cGMP-induced alterations in the cytoskeleton.