Search PubMed⌕ Search

Biomedical subjects

R Franco

Publications and source records attributed to R Franco.

At least 19 recordsLinked to original sources
PubMed2002

Interactions among adenosine deaminase, adenosine A(1) receptors and dopamine D(1) receptors in stably cotransfected fibroblast cells and neurons.

The role of adenosine deaminase in the interactions between adenosine A(1) and dopamine D(1) receptors was studied in a mouse fibroblast cell line stably cotransfected with human D(1) receptor and A(1) receptor cDNAs (A(1)D(1) cells). Confocal laser microscopy analysis showed a high degree of adenosine deaminase immunoreactivity on the membrane of the A(1)D(1) cells but not of the D(1) cells (only cotransfected with human D(1) receptor cDNAs). In double immunolabelling experiments in A(1)D(1) cells and cortical neurons a marked overlap in the distribution of the A(1) receptor and adenosine deaminase immunoreactivities and of the D(1) receptor and adenosine deaminase immunoreactivities was found. Quantitative analysis of A(1)D(1) cells showed that adenosine deaminase immunoreactivity to a large extent colocalizes with A(1) and D(1) receptor immunoreactivity, respectively. The A(1) receptor agonist caused in A(1)D(1) cells and in cortical neurons coaggregation of A(1) receptors and adenosine deaminase, and of D(1) receptors and adenosine deaminase. The A(1) receptor agonist-induced aggregation was blocked by R-deoxycoformycin, an irreversible adenosine deaminase inhibitor. The competitive binding experiments with the D(1) receptor antagonist [(3)H]SCH-23390 showed that the D(1) receptors had a better fit for two binding sites for dopamine, and treatment with the A(1) receptor agonist produced a disappearance of the high-affinity site for dopamine at the D(1) receptor. R-Deoxycoformycin treatment, which has previously been shown to block the interaction between adenosine deaminase and A(1) receptors, and which is crucial for the high-affinity state of the A(1) receptor, also blocked the A(1) receptor agonist-induced loss of high-affinity D(1) receptor binding. The conclusion of the present studies is that the high-affinity state of the A(1) receptor is essential for the A(1) receptor-mediated antagonistic modulation of D(1) receptors and for the A(1) receptor-induced coaggregates of A(1) and adenosine deaminase, and of D(1) and adenosine deaminase. Thus, the confocal experiments indicate that both A(1) and D(1) receptors form agonist-regulated clusters with adenosine deaminase, where the presence of a structurally intact adenosine deaminase bound to A(1) receptors is important for the A(1)-D(1) receptor-receptor interaction at the level of the D(1) receptor recognition.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed2001

Estrogen receptor beta expression in human prostate tissue.

Estrogen receptor subtype beta (ERbeta) is highly expressed in rat prostate epithelium, but its presence in human prostate needs to be confirmed. Here we investigated the expression of ERbeta in five benign (normal and/or hyperplastic) and 10 malignant (Gleasons' score 2-7) prostate tissue specimens using immunohistochemistry. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded tissue sections, using a commercially available ERbeta polyclonal antibody developed against the C-terminal amino acid residue. Nuclear ERbeta expression was found in the nuclei of glandular epithelium of benign prostate tissue specimens; faint nuclear ERbeta positivity was also present in a few stromal cells around normal epithelium. Nuclear ERbeta specific immunostaining was undetectable in all prostate cancer sections.

Animals↗
PubMed2001

Substitution of murine ferrochelatase glutamate-287 with glutamine or alanine leads to porphyrin substrate-bound variants.

Ferrochelatase (EC 4.99.1.1) is the terminal enzyme of the haem biosynthetic pathway and catalyses iron chelation into the protoporphyrin IX ring. Glutamate-287 (E287) of murine mature ferrochelatase is a conserved residue in all known sequences of ferrochelatase, is present at the active site of the enzyme, as inferred from the Bacillus subtilis ferrochelatase three-dimensional structure, and is critical for enzyme activity. Substitution of E287 with either glutamine (Q) or alanine (A) yielded variants with lower enzymic activity than that of the wild-type ferrochelatase and with different absorption spectra from the wild-type enzyme. In contrast to the wild-type enzyme, the absorption spectra of the variants indicate that these enzymes, as purified, contain protoporphyrin IX. Identification and quantification of the porphyrin bound to the E287-directed variants indicate that approx. 80% of the total porphyrin corresponds to protoporphyrin IX. Significantly, rapid stopped-flow experiments of the E287A and E287Q variants demonstrate that reaction with Zn(2+) results in the formation of bound Zn-protoporphyrin IX, indicating that the endogenously bound protoporphyrin IX can be used as a substrate. Taken together, these findings suggest that the structural strain imposed by ferrochelatase on the porphyrin substrate as a critical step in the enzyme catalytic mechanism is also accomplished by the E287A and E287Q variants, but without the release of the product. Thus E287 in murine ferrochelatase appears to be critical for the catalytic process by controlling the release of the product.

Alanine↗
PubMed2001

Metabotropic glutamate 1alpha and adenosine A1 receptors assemble into functionally interacting complexes.

Recently, evidence has emerged that seven transmembrane G protein-coupled receptors may be present as homo- and heteromers in the plasma membrane. Here we describe a new molecular and functional interaction between two functionally unrelated types of G protein-coupled receptors, namely the metabotropic glutamate type 1alpha (mGlu(1alpha) receptor) and the adenosine A1 receptors in cerebellum, primary cortical neurons, and heterologous transfected cells. Co-immunoprecipitation experiments showed a close and subtype-specific interaction between mGlu(1alpha) and A1 receptors in both rat cerebellar synaptosomes and co-transfected HEK-293 cells. By using transiently transfected HEK-293 cells a synergy between mGlu(1alpha) and A1 receptors in receptor-evoked [Ca(2+)](i) signaling has been shown. In primary cultures of cortical neurons we observed a high degree of co-localization of the two receptors, and excitotoxicity experiments in these cultures also indicate that mGlu(1alpha) and A1 receptors are functionally related. Our results provide a molecular basis for adenosine/glutamate receptors cross-talk and open new perspectives for the development of novel agents to treat neuropsychiatric disorders in which abnormal glutamatergic neurotransmission is involved.

Animals↗
PubMed2001

Comodulation of CXCR4 and CD26 in human lymphocytes.

We provide convergent and multiple evidence for a CD26/CXCR4 interaction. Thus, CD26 codistributes with CXCR4, and both coimmunoprecipitate from membranes of T (CD4(+)) and B (CD4(-)) cell lines. Upon induction with stromal cell-derived factor 1alpha (SDF-1alpha), CD26 is cointernalized with CXCR4. CXCR4-mediated down-regulation of CD26 is not induced by antagonists or human immunodeficiency virus (HIV)-1 gp120. SDF-1alpha-mediated down-regulation of CD26 is not blocked by pertussis toxin but does not occur in cells expressing mutant CXCR4 receptors unable to internalize. Codistribution and cointernalization also occurs in peripheral blood lymphocytes. Since CD26 is a cell surface endopeptidase that has the capacity to cleave SDF-1alpha, the CXCR4.CD26 complex is likely a functional unit in which CD26 may directly modulate SDF-1alpha-induced chemotaxis and antiviral capacity. CD26 anchors adenosine deaminase (ADA) to the lymphocyte cell surface, and this interaction is blocked by HIV-1 gp120. Here we demonstrate that gp120 interacts with CD26 and that gp120-mediated disruption of ADA/CD26 interaction is a consequence of a first interaction of gp120 with a domain different from the ADA binding site. SDF-1alpha and gp120 induce the appearance of pseudopodia in which CD26 and CXCR4 colocalize and in which ADA is not present. The physical association of CXCR4 and CD26, direct or part of a supramolecular structure, suggests a role on the function of the immune system and the pathophysiology of HIV infection.

Adenosine Deaminase↗
PubMed2001

Medullary thyroid cancer, papillary thyroid microcarcinoma and Graves' disease: an unusual clinical coexistence.

We describe the unusual case of a Caucasian woman who had a diagnosis of medullary thyroid cancer and papillary microcarcinoma 5 years after a diagnosis of Graves' disease. The patient came to our observation for recurrence of hyperthyroidism. An ultrasound scan revealed diffuse thyroid enlargement with a nodule, recently increased in size. The serum CT and carcinoembrional antigen were elevated, and the fine-needle aspiration cytology with immunocytochemical analysis for CT was suggestive for medullary thyroid carcinoma. The nodular lesion showed intense 111In-pentetreotide uptake, whereas total body scintigraphy with the same tracer and with Thallium-201, 99mTc (V) dimercaptosuccinic acid was negative for lymph node and distant metastasis. The histological examination of thyroidectomy specimens confirmed the diagnosis of medullary thyroid cancer, showing a lymphocytic intratumoral infiltration. The histological analysis of the controlateral lobe showed an occult papillary microcarcinoma. Medullary thyroid carcinoma and papillary microcarcinoma showed intense staining with policlonal anti-RET antibodies, although genetic analysis was negative for RET mutations most frequently involved in familial and sporadic medullary thyroid carcinomas. Possible implications about the coexistence of the 3 thyroid diseases are discussed.

Adult↗
PubMed2001

Synchronous primary adenocarcinoma and adenosquamous carcinoma of the esophagus.

Multiple malignant esophageal tumors of the same cell type are described. In the esophageal mucosa, widespread carcinomatous transformation may be observed and multicentric invasive squamous cell carcinomas may develop. The concomitance of two independent esophageal malignant neoplasms of different epithelial histogenesis is uncommon. Synchronous adenocarcinoma and squamous cell carcinoma of the esophagus is reported. Adenosquamous carcinoma of the esophagus is a rare tumor. Adenocarcinoma of the esophagus represents 10% of esophageal cancer. We report a case of a synchronous primary invasive adenosquamous carcinoma and adenocarcinoma of the esophagus. Both tumors were demonstrated radiographically. The peculiarity of this neoplastic association and the importance of complete radiographic esophageal evaluation in patients with one obvious obstructing tumor of the esophagus are emphasized.

Adenocarcinoma↗
PubMed2001

Evidence for two mechanisms of amino acid osmolyte release from hippocampal slices.

A 30% decrease in osmolarity stimulated 3H-taurine, 3H-GABA and glutamate (followed as 3H-D-aspartate) efflux from rat hippocampal slices. 3H-taurine efflux was activated rapidly but inactivated slowly. It was decreased markedly by 100 microM 5-nitro-(3-phenylpropylamino)benzoic acid (NPPB) and 600 microM niflumic acid and inhibited strongly by tyrphostins AG18, AG879 and AG112 (25-100 microM), suggesting a tyrosine kinase-mediated mechanism. Hyposmolarity activated the mitogen-activated protein kinases (MAPK) extracellular-signal-related kinase-1/2 (ERK1/ERK2) and p38, but blockade of this reaction did not affect 3H-taurine efflux. Hyposmosis also activated phosphatidylinositide 3-kinase (PI3K) and its prevention by wortmannin (100 nM) essentially abolished 3H-taurine efflux. 3H-taurine efflux was insensitive to the protein kinase C (PKC) blocker chelerythrine (2.5 microM) or to cytochalasin E (3 microM). The release of 3H-GABA and 3H-D-aspartate occurred by a different mechanism, characterized by rapid activation and inactivation, insensitivity to NPPB, niflumic acid, tyrphostins or wortmannin. 3H-GABA and 3H-D-aspartate efflux was not due to external [NaCl] decrease, cytosolic Ca2+ increase or depolarization, or to reverse operation of the carrier. This novel mechanism of amino acid release may be mediated by Ca2+-independent exocytosis and modulated by PKC and actin cytoskeleton disruption, as suggested by its inhibition by chelerythrine and potentiation by 100 nM phorbol-12-myristate-13 acetate (PMA) and cytochalasin E. GABA and glutamate osmosensitive efflux may explain the hyposmolarity-elicited increase in amplitude of inhibitory and excitatory postsynaptic potentials in hippocampal slices as well as the hyperexcitability associated with hyponatraemia.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2001

Expression of PCNA in the testis of the lizard, Podarcis s. sicula: an endogenous molecular marker of mitotic germinal epithelium proliferation.

Proliferating cell nuclear antigen (PCNA) plays an essential role in nucleic acid metabolism as a component of the replication and repair machinery. This protein encircles DNA and can slide bidirectionally along the duplex binding to DNA polymerase delta and epsilon. It is well known that PCNA interacts with proteins involved in the cell cycle. The PCNA interactions with different cellular proteins and the importance of these interactions are discussed. To examine mitotic germinal epithelium proliferation during annual discontinuous spermatogenesis in the lizard Podarcis s. sicula, temporal and the spatial PCNA expression were investigated, and provide a useful endogenous molecular marker.

Animals↗
PubMed2001

Variations of proline-rich kinase Pyk2 expression correlate with prostate cancer progression.

Proline-rich kinase 2 (Pyk2), also known as CAKbeta (cell adhesion kinase beta), is a cytoplasmic tyrosine kinase that is structurally related to focal adhesion kinase. Pyk2 is expressed in different cell types including brain cells, fibroblasts, platelets, and other hemopoietic cells. Pyk2 is rapidly tyrosine phosphorylated in response to diverse extracellular signals acting via different post receptor pathways. We have investigated whether this protein kinase is functionally expressed in normal and neoplastic prostate tissues. In this study, we demonstrate that Pyk2 is expressed only in normal epithelial prostate tissue and in benign prostatic hyperplasia, whereas its expression progressively declines with an increasing grade of malignancy of prostate cancer.

Enzyme Activation↗
PubMed2001

Cutaneous presentation of follicular lymphomas.

The description of primary cutaneous follicular lymphoma has raised interest in the differential diagnosis of this versus disseminated follicular lymphoma involving the skin. We report here on four cases of Stage IV follicular lymphoma, diagnosed in skin biopsy, in which cutaneous lesion was the most noticeable feature of clinical presentation. In all cases, the morphological features were superimposed over typical nodal follicular lymphoma. Apart from classic B-cell markers, they were characterized by CD10 and bcl6 positivity, markers of follicle germinal center cells; and bcl2 expression, with a corresponding t(14;18) translocation in three of three cases examined. In all four cases, bone marrow study and clinical staging revealed disease that had disseminated since diagnosis. Follow-up showed relapsing cutaneous and nodal disease in two cases. The only difference observed with a control group of 10 cases of primary cutaneous follicular lymphoma was the absence in this group of t(14; 18). Disseminated classical follicular lymphoma has to be considered in the differential diagnosis of follicular lymphoma presenting in the skin. This series of cases suggests that the presence of t(14;18) could imply the existence of disease that has disseminated beyond the skin and that cases harboring this translocation could be candidates for systemic polychemotherapy.

Adult↗
PubMed2001

Cutaneous follicular B-cell lymphoma: description of a series of 18 cases.

The lack of precise and homogeneous criteria for the recognition of primary cutaneous follicular lymphoma has hindered gaining data on the frequency and clinical and molecular features of this entity. In the course of a review of a series of primary cutaneous lymphoma from different Spanish hospitals, we collected a series of 18 cases of primary cutaneous follicular lymphoma and analyzed its clinical, morphologic, and biologic characteristics. In this review only cases with a follicular pattern of growth, germinal center cytology, and restriction to the skin in a minimum follow-up of 6 months have been included. Cases of primary cutaneous follicular lymphoma were characterized by the expression of classic markers of the germinal center, such as bcl6, CD10, and the presence of aggregates of follicular dendritic cells. They frequently express bcl2 protein, although classical t(14;18) was not found in any of the cases analyzed. Analysis of the bcl6 noncoding first exon showed somatic mutations in two of four cases analyzed, as would be expected in lymphoma deriving from the germinal center. Clinically, most cases showed initial involvement of the head and neck, with relapses in eight cases (involving the skin in five cases, both skin and lymph node in two cases, and lymph node in one case). No death attributable to the tumor was recorded. These data seem to imply that follicular lymphoma may present initially in the skin, lacking the characteristic t(14;18) and having a relatively indolent course. Recognition of these tumors and elucidation of their molecular alterations could lead to properly adapted staging and treatment protocols for these patients.

Adult↗
PubMed2001

Microvascular coagulopathy and disseminated intravascular coagulation.

OBJECTIVE: To review the dual characteristics of disseminated intravascular coagulation (DIC), as both a contributor to multiple organ failure as well as a symptom of severe underlying disease associated with systemic vascular changes. DATA SOURCES: Published literature data and unpublished results from the authors. DATA SUMMARY: Clinical and experimental studies strongly suggest that DIC contributes to multiple organ failure and death in patients with severe systemic disorders such as sepsis. DIC is evoked by systemic cytokine activity, and the inflammatory response aggravates vascular permeability, inflammation, and cell damage in tissues. In addition to intravascular fibrin formation, thrombin and fibrin generation in tissues is also an important aspect of DIC. An example of DIC at the organ level is adult respiratory distress syndrome, where fibrin in the lung is a characteristic feature. Intravascular fibrin formation and occlusion may elicit a hypoxic response with induction of hypoxia related transcription factors. The resulting ischemic preconditioning may offer protective effects to the involved organ(s). CONCLUSIONS: Overall, the beneficial or harmful effects of activated coagulation and fibrin formation for organ pathology and recovery from DIC remain to be explored. This may be a critical element in the assessment of ischemia-reperfusion effects of specific anticoagulant therapy.

Anticoagulants↗
PubMed2001

DNA ploidy and cyclin D1 expression in basal cell carcinoma of the head and neck.

Basal cell carcinomas (BCCs) may be subdivided into primary with a favorable biologic course (BCC1) and recurrent and/or metastatic (BCC2). No clear association between primary tumor location, histologic subtype, or other clinicopathologic variables and predisposition for BCC2 has been found. Histopathologic criteria are limited for prognostication. To identify prognostic factors useful for planning therapy, we studied cyclin D1 immunohistochemical expression, DNA ploidy, and epiluminescence light microscopic (ELM) patterns in 60 cases of BCC (30 BCC1 and 30 BCC2) in the head and neck region, half of which were hyperpigmented. Cyclin D1 was absent in 27 cases, expressed at low level in 4 cases, and overexpressed in 30 cases. Seven BCCs were euploid, 28 exhibited a mixed cellular population, and 25 were aneuploid. Among aneuploid tumors, hypodiploidy was found in 12. Among the 30 pigmented carcinomas, only 15 showed a typical ELM pattern. No association between pigmentation and more aggressive biologic behavior of BCC was found. These results and follow-up data seem to indicate that an unfavorable outcome can be predicted by hyperexpression of cyclin D1, aneuploidy, and an atypical ELM pattern for pigmented cases. A definite hypodiploid peak was associated with worse prognosis. The analysis of cyclin D1 expression and DNA ploidy may help identify BCC with an aggressive phenotype and a poor clinical outcome.

Basal Cell Carcinoma↗
PubMed2001

P53 and hMSH2 expression in basal cell carcinomas and malignant melanomas from photoexposed areas of head and neck region.

Ultraviolet (UV) radiation plays a pivotal role in skin damage and photocarcinogenesis. The basic mechanism of phototoxicity lies in DNA damage, and involves mutation of tumor suppressor genes, oncogenes and genes directly involved in the control of the stability of genome, such as the mismatch repair (MR) genes. The goal of this study was to evaluate the role of p53 and hMSH2 in the UV-related carcinogenetic process. An immunohistochemical study for p53 and hMSH2 was performed in a series of 43 basal cell carcinomas (BCC) and 60 melanomas (MM) from photoexposed areas of head and neck region, comparing the findings with follow-up. A deregulated p53 expression characterized less differentiated, more aggressive BCC (BCC2) but not the well-differentiated ones (BCC1). The hMSH2 protein was present, though expressed at varying levels, in 18 out of 21 BCC1 cases and in 4 out of 22 BCC2. In the remaining 3 cases of BCC1 and 18 cases of BCC2, a complete absence of hMSH2 expression was found, correlating directly with the presence of recurrence and/or death of the disease in case of melanoma (p<0.05). Overall, the expression of hMSH2 correlated inversely with the p53 overexpression (p<0.01). In MM, p53 was found overexpressed in 81.6% of the cases, and this correlated positively with the level of infiltration and with the presence of relapses (p<0.01) or metastasis (p<0.01) and inversely with the disease-free interval (p<0.05). These results are in agreement with the reported association between p53 deregulation and a more aggressive cancer phenotype. The evaluation of the expression of p53 and hMSH2 could improve the management of patients with BCC and MM, and could have a role also in the evaluation of the early cutaneous photo-inducted damage, contributing to the identification of presymptomatic patients predisposed to the development of UV-related new skin tumors, who could become candidates for chemoprevention trials.

Adult↗
PubMed2001

Adenosine A2B receptors behave as an alternative anchoring protein for cell surface adenosine deaminase in lymphocytes and cultured cells.

Adenosine deaminase (ADA) is an enzyme of the purine metabolism that has been largely considered to be cytosolic. Recently, it has been demonstrated that the enzyme appears on the surface of lymphocytes where it interacts with the T-cell activation antigen CD26. ADA also appears on the surface of nonlymphoid cells anchored to adenosine A1 receptors. Here it is demonstrated that cell surface ADA in ADA+/CD26- T lymphocytes anchors to adenosine receptors of the A2B subtype (A2BR). An interaction between A2BR and cell surface ADA has been demonstrated in transfected Chinese hamster ovary cells and Jurkat J32 T lymphocytes. This has been proved by coimmunoprecipitation, binding of exogenous ADA to A2BR+ cells, and coimmunolocalization. The specificity of the interaction has also been demonstrated by the lack of interaction with other members of the G protein-coupled receptor superfamily. Binding of ADA to A2BR increases the affinity of the agonist 5'-N-ethylcarboxamidoadenosine and cAMP production. This effect occurs even when ADA devoid of enzyme activity is used. Therefore, in lymphocytes, cell surface ADA, apart from degrading extracellular adenosine, regulates those actions of adenosine that are mediated via adenosine receptors of the A2B subtype.

Adenosine Deaminase↗
PubMed2001

Involvement of caveolin in ligand-induced recruitment and internalization of A(1) adenosine receptor and adenosine deaminase in an epithelial cell line.

Chronic exposure of A(1) adenosine receptors (A(1)R) to A(1)R agonists leads to activation, phosphorylation, desensitization, and internalization to intracellular compartments of the receptor. Desensitization and internalization of A(1)R is modulated by adenosine deaminase (ADA), an enzyme that regulates the extracellular concentration of adenosine. ADA interacts with A(1)R on the cell surface of the smooth muscle cell line DDT1 MF-2, and both proteins are internalized following agonist stimulation of the receptor. The mechanism involved in A(1)R and ADA internalization upon agonist exposure is poorly understood in epithelial cells. In this report, we show that A(1)R and ADA interact in LLC-PK(1) epithelial cells. Exposure of LLC-PK(1) cells to A(1)R agonists induces aggregation of A(1)R and ADA on the cell surface and their translocation to intracellular compartments. Biochemical and cell biology assays were used to characterize the intracellular vesicles containing both proteins after agonist treatment. A(1)R and ADA colocalized together with the rafts marker protein caveolin. Filipin, a sterol-binding agent that disrupts rafts (small microdomains of the plasma membrane), was able to inhibit A(1)R internalization. In contrast, acid treatment of the cells, which disrupts internalization via clathrin-coated vesicles, did not inhibit agonist-stimulated A(1)R internalization. We demonstrated that A(1)R agonist N(6)-(R)-phenylisopropyl adenosine promotes the translocation of A(1)R into low-density gradient fractions containing caveolin. Furthermore, a direct interaction of the C-terminal domain of A(1)R with caveolin-1 was demonstrated by pull down experiments. These results indicate that A(1)R and ADA form a stable complex in the cell surface of LLC-PK(1) cells and that agonist-induced internalization of the A(1) adenosine receptor and ADA is mediated by clathrin-independent endocytosis.

Adenosine Deaminase↗