Correlation between Different Psychological Variables in Women with Fibromyalgia with Symptoms of Neurogenic Inflammation: A Cross-Sectional Study.

Fibromyalgia (FM) is a chronic pain syndrome hypothesized to arise from a state of neurogenic inflammation. Mechanisms responsible for pain, as well as psychological variables, are typically altered in this condition. The main objective of this research was to explore somatosensory and psychological alterations in women with FM. The secondary objective was to carry out a secondary analysis to correlate the different variables studied and delve into the influences between them. The relationship between different psychological variables in fibromyalgia is not clear in the previous scientific literature. Forty-four individuals participated, of which twenty-two were controls and twenty-two were women with fibromyalgia. The main outcome measures were the Numeric Pain Rating Scale, Fibromyalgia Impact Questionnaire, pressure pain threshold, conditioned pain modulation, anxiety and depression symptoms, catastrophizing and kinesiophobia cognitions. The main analysis showed that there is a moderate correlation between the psychological variables of depression and fear of movement and the ability to modulate pain. There is also a moderately inverse correlation between pain catastrophizing cognitions and pain intensity/disability. Multiple moderate and strong correlations were found among the various psychological variables studied. FM patients exhibit somatosensory alterations alongside negative psychological symptoms that influence the experience of pain, and they may perpetuate the state of neurogenic inflammation.

Apolipoprotein E (ApoE) ε4 Genotype (ApoE rs429358-ApoE rs7412 Polymorphisms) Is Not Associated with Long COVID Symptoms in Previously Hospitalized COVID-19 Survivors.

The role of genetics as a predisposing factor related to an increased risk of developing long COVID symptomatology is under debate. The aim of the current secondary analysis was to identify the association between the Apolipoprotein E (ApoE) gene, a gene affecting cholesterol metabolism and previously associated with a higher risk of SARS-CoV-2 infection and COVID-19 severity, and the development of long COVID in a cohort of individuals who had been hospitalized by SARS-CoV-2 infection. Unstimulated whole saliva samples were collected from 287 previously hospitalized COVID-19 survivors. Three genotypes of the ApoE gene (ApoE ε2, ε3, ε4) were obtained based on the combination of ApoE rs429358 and ApoE rs7412 polymorphisms. Participants were asked to self-report the presence of any post-COVID symptom in a face-to-face interview at 17.8 ± 5.2 months after hospital discharge and medical records were obtained. Each participant reported 3.0 (1.9) post-COVID symptoms. Overall, no significant differences in long COVID symptoms were observed depending on the ApoE genotype (ApoE ε2, ApoE ε3, ApoE ε4). The presence of the ApoE ε4 genotype, albeit associated with a higher risk of SARS-CoV-2 infection and COVID-19 severity, did not appear to predispose for the presence of long COVID in our cohort of previously hospitalized COVID-19 survivors.

Working memory dysfunction in fibromyalgia is associated with genotypes of the catechol- O-methyltransferase gene: an event-related potential study.

Recent findings have associated different COMT genotypes with working memory capacity in patients with fibromyalgia. Although it is thought that the COMT gene may influence neural correlates (P2 and P3 ERP components) underlying working memory impairment in this chronic-pain syndrome, it has not yet been explored. Therefore, the aim of the present research was to investigate the potential effect of the COMT gene in fibromyalgia patients on ERP working memory indices (P2 and P3 components). For this purpose, 102 participants (51 patients and 51 healthy control participants) took part in the experiment. Event-related potentials and behavioral responses were recorded while participants performed a spatial n-back task. Participants had to decide if the stimulus coincided or not in the same location as the one presented one (1-back condition) or two (2-back condition) trials before. Genotypes of the COMT gene were determined through a saliva sample from all participants. Present results significantly showed lower working memory performance (p < 0.05) in patients with fibromyalgia as compared to control participants (higher rate of errors and slower reaction times). At neural level, we found that patients exhibited enhanced frontocentral and parieto-occipital P2 amplitudes compared to control participants (p < 0.05). Interestingly, we also observed that only fibromyalgia patients carrying the Val/Val genotype of the COMT gene showed higher frontocentral P2 amplitudes than control participants (p < 0.05). Current results (behavioral outcomes and P2 amplitudes) confirmed the presence of an alteration in working memory functioning in fibromyalgia. The enhancement of frontocentral P2 could be reflecting that these patients would manifest an inefficient way of activating executive attention processes, in carriers of the Val/Val genotype of COMT. To our knowledge, the present findings are the first linking neural indices of working memory dysfunctions and COMT genotypes in fibromyalgia. Applying a subgroup of patient's strategy based on this genetic marker could be useful to establish more tailored therapeutical approaches.

Genetic Association between ACE2 (rs2285666 and rs2074192) and TMPRSS2 (rs12329760 and rs2070788) Polymorphisms with Post-COVID Symptoms in Previously Hospitalized COVID-19 Survivors.

The aim of the study was to identify the association between four selected COVID-19 polymorphisms of ACE2 and TMPRSS2 receptors genes with the presence of long-COVID symptomatology in COVID-19 survivors. These genes were selected as they associate with the entry of the SARS-CoV-2 virus into the cells, so polymorphisms could be important for the prognoses of long-COVID symptoms. Two hundred and ninety-three (n = 293, 49.5% female, mean age: 55.6 ± 12.9 years) individuals who had been previously hospitalized due to COVID-19 were included. Three potential genotypes of the following single nucleotide polymorphisms (SNPs) were obtained from non-stimulated saliva samples of participants: ACE2 (rs2285666), ACE2 (rs2074192), TMPRSS2 (rs12329760), TMPRSS2 (rs2070788). Participants were asked to self-report the presence of any post-COVID defined as a symptom that started no later than one month after SARS-CoV-2 acute infection and whether the symptom persisted at the time of the study. At the time of the study (mean: 17.8, SD: 5.2 months after hospital discharge), 87.7% patients reported at least one symptom. Fatigue (62.8%), pain (39.9%) or memory loss (32.1%) were the most prevalent post-COVID symptoms. Overall, no differences in long-COVID symptoms were dependent on ACE2 rs2285666, ACE2 rs2074192, TMPRSS2 rs12329760, or TMPRSS2 rs2070788 genotypes. The four SNPs assessed, albeit previously associated with COVID-19 severity, do not predispose for developing long-COVID symptoms in people who were previously hospitalized due to COVID-19 during the first wave of the pandemic.

Post-COVID Pain Is Not Associated with Inflammatory Polymorphisms in People Who Had Been Hospitalized by COVID-19.

Our aim was to assess the association between four inflammatory polymorphisms with the development of post-COVID pain and to associate these polymorphisms with the clinical pain phenotype in individuals who had been hospitalized by COVID-19. Three potential genotypes of IL-6 (rs1800796), IL-10 (rs1800896), TNF-α (rs1800629), and IFITM3 (rs12252) single nucleotide polymorphisms (SNPs) were obtained from no-stimulated saliva samples from 293 (49.5% female, mean age: 55.6 ± 12.9 years) previously hospitalized COVID-19 survivors by polymerase chain reactions. Pain phenotyping consisted of the evaluation of pain features, sensitization-associated symptoms, anxiety levels, depressive levels, sleep quality, catastrophizing, and kinesiophobia levels in patients with post-COVID pain. Analyses were conducted to associate clinical features with genotypes. One hundred and seventeen (39.9%) patients experienced post-COVID pain 17.8 ± 5.2 months after hospital discharge. No significant differences in the distribution of the genotype variants of any SNPs were identified between COVID-19 survivors with and without post-COVID pain (all, p > 0.47). Similarly, the clinical pain phenotype was not significantly different between patients with and without post-COVID pain since no differences in any variable were observed for any SNPs. In conclusion, four SNPs associated with inflammatory and immune responses did not appear to be associated with post-COVID pain in previously hospitalized COVID-19 survivors. Further, neither of the SNPs were involved in the phenotyping features of post-COVID pain.

Fear of pain moderates the relationship between self-reported fatigue and methionine allele of catechol-O-methyltransferase gene in patients with fibromyalgia.

Previous research has shown a consistent association among genetic factors, psychological symptoms and pain associated with fibromyalgia. However, how these symptoms interact to moderate genetic factors in fibromyalgia has rarely been studied to date. The present research investigates whether psychological symptoms can moderate the effects of catechol-O-methyltransferase on pain and fatigue. A total of 108 women diagnosed with fibromyalgia and 77 healthy control participants took part in the study. Pain, fatigue, and psychological symptoms (anxiety, depression, pain catastrophizing, fear of pain and fear of movement) were measured by self-report questionnaires. Two types of statistical analyses were performed; the first was undertaken to explore the influences of COMT genotypes on clinical symptoms by comparing patients with fibromyalgia and healthy controls. In the second analysis, moderation analyses to explore the role of psychological symptoms as potential factors that moderate the relationship between pain/fatigue and COMT genotypes were performed. The main results indicated that patients carrying the Met/Met genotype reported significantly higher levels of fatigue than heterozygote carriers (i.e., Met/Val genotype) and higher levels of fatigue, but not significantly different, than Val homozygote carriers. Among patients with fibromyalgia carrying methionine alleles (i.e., Met/Met + Met/Val carriers), only those who scored high on medical fear of pain, experienced an intensified feeling of fatigue. Thus, the present research suggests that fear of pain, as a psychological symptom frequently described in fibromyalgia may act as a moderating factor in the relationship between the Met allele of the COMT gene and the increase or decrease in self-reported fatigue. Although further research with wider patient samples is needed to confirm the present findings, these results point out that the use of psychological interventions focused on affective symptomatology might be a useful tool to reduce the severity of fibromyalgia.

Effects of COMT Genotypes on Working Memory Performance in Fibromyalgia Patients.

Growing research has reported the presence of a clear impairment of working memory functioning in fibromyalgia. Although different genetic factors involving dopamine availability (i.e, the COMT gene) have been associated with the more severe presentation of key symptoms in fibromyalgia, scientific evidence regarding the influence of COMT genotypes on cognitive impairment in these patients is still lacking. To this end, 167 participants took part in the present investigation. Working memory performance was assessed by the application of the SST (Spatial Span Test) and LNST (Letter and Number Sequence Test) belonging to the Weschler Memory Scale III. Significant working memory impairment was shown by the fibromyalgia patients. Remarkably, our results suggest that performance according to different working memory measures might be influenced by different genotypes of the COMT gene. Specifically, fibromyalgia patients carrying the Val/Val genotype exhibited significantly worse outcomes for the span of SST backward, SST backward score, SST total score and the Working Memory Index (WMI) than the Val/Val healthy carriers. Furthermore, the Val/Val patients performed worse on the SST backward and SST score than heterozygotes. Our findings are the first to show a link between the COMT gene and working memory dysfunction in fibromyalgia, supporting the idea that higher COMT enzyme activity would contribute to more severe working memory impairment in fibromyalgia.

Influence of the Topographic Vascular Distribution of the Face on Dermal Filler Accidents.

OBJECTIVE: Dermal fillers are an important tool in the field of aesthetic dermatology. Fillers are relatively noninvasive and easy to use but are not free of secondary complications. The main complications are vascular and are due to either the compression of an artery or the direct introduction of the product into the arterial lumen. The aim of this study is to provide an overview of the vascular territories of the face to avoid many possible complications when using facial fillings. Anatomical localization of the main arterial supply to the face has been described to assess the risk of vascular injury. METHODS: The authors dissected 17 hemifaces of embalmed adult cadavers that had previously been injected, through the common carotid artery, with latex containing a red dye. RESULTS: A topographic distribution was generated by facial regions following a clinical approach from where the facial fillings were placed and related to the pathways of the arteries. Following these criteria, we established 8 topographic regions (I-VIII) that indicate the main vascular problems of each of these regions. Detailed anatomical localizations of the main arteries in these topographic regions of the face and their relationships are described. CONCLUSIONS: The highest index of vascular lesions and especially visual alterations occurred for fillings of the upper third of the face. To prevent and avoid this type of lesion, it is advisable to avoid, as much as possible, treatments with filling materials in the upper third of the face, mainly including the glabellar and nasal region (III) and supraorbital region (VIII).

Changes in the cellular microRNA profile by the intracellular expression of HIV-1 Tat regulator: A potential mechanism for resistance to apoptosis and impaired proliferation in HIV-1 infected CD4+ T cells.

HIV-1 induces changes in the miRNA expression profile of infected CD4+ T cells that could improve viral replication. HIV-1 regulator Tat modifies the cellular gene expression and has been appointed as an RNA silencing suppressor. Tat is a 101-residue protein codified by two exons that regulates the elongation of viral transcripts. The first exon of Tat (amino acids 1-72) forms the transcriptionally active protein Tat72, but the presence of the second exon (amino acids 73-101) results in a more competent regulatory protein (Tat101) with additional functions. Intracellular, full-length Tat101 induces functional and morphological changes in CD4+ T cells that contribute to HIV-1 pathogenesis such as delay in T-cell proliferation and protection against FasL-mediated apoptosis. But the precise mechanism by which Tat produces these changes remains unknown. We analyzed how the stable expression of intracellular Tat101 and Tat72 modified the miRNA expression profile in Jurkat cells and if this correlated with changes in apoptotic pathways and cell cycle observed in Tat-expressing cells. Specifically, the enhanced expression of hsa-miR-21 and hsa-miR-222 in Jurkat-Tat101 cells was associated with the reduced expression of target mRNAs encoding proteins related to apoptosis and cell cycle such as PTEN, PDCD4 and CDKN1B. We developed Jurkat cells with stable expression of hsa-miR-21 or hsa-miR-222 and observed a similar pattern to Jurkat-Tat101 in resistance to FasL-mediated apoptosis, cell cycle arrest in G2/M and altered cell morphology. Consequently, upregulation of hsa-miR-21 and hsa-miR-222 by Tat may contribute to protect against apoptosis and to anergy observed in HIV-infected CD4+ T cells.

Gallus gallus orthologous to human alpha-dystroglycanopathies candidate genes: Gene expression and characterization during chicken embryogenesis.

Alpha-dystroglycanopathies are a heterogenic group of human rare diseases that have in common defects of α-dystroglycan O-glycosylation. These congenital disorders share common features as muscular dystrophy, malformations on central nervous system and more rarely altered ocular development, as well as mutations on a set of candidate genes involved on those syndromes. Severity of the syndromes is variable, appearing Walker-Warburg as the most severe where mutations at protein O-mannosyl transferases POMT1 and POMT2 genes are frequently described. When studying the lack of MmPomt1 in mouse embryonic development, as a murine model of Walker-Warburg syndrome, MmPomt1 null phenotype was lethal because Reitchert's membrane fails during embryonic development. Here, we report gene expression from Gallus gallus orthologous genes to human candidates on alpha-dystroglycanopathies POMT1, POMT2, POMGnT1, FKTN, FKRP and LARGE, making special emphasis in expression and localization of GgPomt1. Results obtained by quantitative RT-PCR, western-blot and immunochemistry revealed close gene expression patterns among human and chicken at key tissues affected during development when suffering an alpha-dystroglycanopathy, leading us to stand chicken as a useful animal model for molecular characterization of glycosyltransferases involved in the O-glycosylation of α-Dystroglycan and its role in embryonic development.

Brain correlates of cognitive inhibition in fibromyalgia: emotional intrusion of symptom-related words.

Evidence coming from neuropsychological studies has showed the presence of cognitive alterations in fibromyalgia. Such dysfunctions are especially remarkable when the set in motion of executive control processes, such as inhibition, is required to perform successfully; however, neural data related to these mechanisms are very scarce. Present study tried to characterize cognitive inhibition mechanisms, as part of the attentional control functions, in patients with fibromyalgia. Participants (two groups: fibromyalgia patients and healthy control participants) were asked to perform in an emotional Stroop task while event-related potentials (ERP) were recorded. Four different emotional interference conditions were created: fibromyalgia symptom-related words, arousing-negative, arousing-positive and neutral words. Brain activity and behavioral data were analyzed. Principal component analyses were employed to reliably define ERP components along with a source-estimation technique. Symptom-related words elicited greater frontal P450 amplitudes and enhanced activation within right inferior frontal gyrus as compared to the rest of stimuli. This effect was only true for the fibromyalgia group. Behavioral contrasts, however, did not produce significant differences. Scalp and source estimation findings suggest the presence of a specific difficulty in cognitive inhibition in fibromyalgia patients (under conditions intimately linked with the core concerns of their disease). Data point to the involvement of right inferior frontal cortices in this inefficient mechanism, which might cause an enhanced and dysfunctional effort of processing to achieve only a comparable performance to healthy people. Implications of these results are discussed. Nevertheless, further investigations are needed to better understand dysfunctional cognition in fibromyalgia.

The presence of HIV-1 Tat protein second exon delays fas protein-mediated apoptosis in CD4+ T lymphocytes: a potential mechanism for persistent viral production.

HIV-1 replication is efficiently controlled by the regulator protein Tat (101 amino acids) and codified by two exons, although the first exon (1-72 amino acids) is sufficient for this process. Tat can be released to the extracellular medium, acting as a soluble pro-apoptotic factor in neighboring cells. However, HIV-1-infected CD4(+) T lymphocytes show a higher resistance to apoptosis. We observed that the intracellular expression of Tat delayed FasL-mediated apoptosis in both peripheral blood lymphocytes and Jurkat cells, as it is an essential pathway to control T cell homeostasis during immune activation. Jurkat-Tat cells showed impairment in the activation of caspase-8, deficient release of mitochondrial cytochrome c, and delayed activation of both caspase-9 and -3. This protection was due to a profound deregulation of proteins that stabilized the mitochondrial membrane integrity, such as heat shock proteins, prohibitin, or nucleophosmin, as well as to the up-regulation of NF-κB-dependent anti-apoptotic proteins, such as BCL2, c-FLIPS, XIAP, and C-IAP2. These effects were observed in Jurkat expressing full-length Tat (Jurkat-Tat101) but not in Jurkat expressing the first exon of Tat (Jurkat-Tat72), proving that the second exon, and particularly the NF-κB-related motif ESKKKVE, was necessary for Tat-mediated protection against FasL apoptosis. Accordingly, the protection exerted by Tat was independent of its function as a regulator of both viral transcription and elongation. Moreover, these data proved that HIV-1 could have developed strategies to delay FasL-mediated apoptosis in infected CD4(+) T lymphocytes through the expression of Tat, thus favoring the persistent replication of HIV-1 in infected T cells.

Protein kinase Ctheta is a specific target for inhibition of the HIV type 1 replication in CD4+ T lymphocytes.

Integration of HIV-1 genome in CD4(+) T cells produces latent reservoirs with long half-life that impedes the eradication of the infection. Control of viral replication is essential to reduce the size of latent reservoirs, mainly during primary infection when HIV-1 infects CD4(+) T cells massively. The addition of immunosuppressive agents to highly active antiretroviral therapy during primary infection would suppress HIV-1 replication by limiting T cell activation, but these agents show potential risk for causing lymphoproliferative disorders. Selective inhibition of PKC, crucial for T cell function, would limit T cell activation and HIV-1 replication without causing general immunosuppression due to PKC being mostly expressed in T cells. Accordingly, the effect of rottlerin, a dose-dependent PKC inhibitor, on HIV-1 replication was analyzed in T cells. Rottlerin was able to reduce HIV-1 replication more than 20-fold in MT-2 (IC(50) = 5.2 µM) and Jurkat (IC(50) = 2.2 µM) cells and more than 4-fold in peripheral blood lymphocytes (IC(50) = 4.4 µM). Selective inhibition of PKC, but not PKCδ or -ζ, was observed at <6.0 µM, decreasing the phosphorylation at residue Thr(538) on the kinase catalytic domain activation loop and avoiding PKC translocation to the lipid rafts. Consequently, the main effector at the end of PKC pathway, NF-κB, was repressed. Rottlerin also caused a significant inhibition of HIV-1 integration. Recently, several specific PKC inhibitors have been designed for the treatment of autoimmune diseases. Using these inhibitors in combination with highly active antiretroviral therapy during primary infection could be helpful to avoid massive viral infection and replication from infected CD4(+) T cells, reducing the reservoir size at early stages of the infection.

Characterization of the BH3 protein Bmf in Gallus gallus: identification of a novel chicken-specific isoform.

Bmf is a proapoptotic member of the BH3-only subgroup of Bcl-2 family proteins, which is associated to myosin V motors by binding to the dynein light chain 2 (DLC2). It acts as a sentinel detecting intracellular damages on the main cytoskeletal structures. The cloning and characterization of the chicken (Gallus gallus) Bmf cDNA and splicing variant is described in this report. The Bmf cDNA was amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) using oligonucleotide primers derived from in silico sequences. The chicken Bmf cDNA encodes a protein of 193 amino acids, showing homology to mammalian Bmf proteins. A splicing variant of the chicken Bmf (Bmf(S), short isoform of Bmf) coding a protein of 118 amino acids was also identified. This is the first Bmf isoform identified so far which lacks the DLC2-binding domain although retaining the BH3 domain. Both chicken Bmf isoforms induced apoptosis 24 h after transfection in MCF7 and HeLa cell lines, but chicken Bmf(S) exhibits a higher proapoptotic activity. In addition, mRNA expression analysis showed that chicken Bmf transcription is ubiquitous in all embryo developmental stages, suggesting a role for this protein in the control of the development process.

Positive correlation between the expression of X-chromosome RBM genes (RBMX, RBM3, RBM10) and the proapoptotic Bax gene in human breast cancer.

In a recent report, it has been postulated that the ubiquitous RBM proteins might constitute a novel family of apoptosis modulators. We measured the expression of the X-chromosome RBM genes (RBMX, RBM3, and RBM10) in 122 breast cancers by means of differential RT-PCR. Using the same method, we also studied the expression of the apoptosis-related genes Bcl-2 and Bax. Markers of hormone dependence (estrogen and progesterone receptors), proliferation (Ki67 and DNA-ploidy), angiogenesis (VEGF and CD105), as well as oncogene (c-erb-B2), and tumor suppressor gene (p53) expression were also analyzed. The expression of all X-chromosome RBM genes was significantly associated with the expression of the proapoptotic Bax gene (RBMX, P=0.039; RBM3, P<0.001; RBM10 large variant, P<0.001; RBM10 small variant, P<0.001). Furthermore, the expression of both RBM10 variants was significantly associated with the expression of the VEGF gene (large variant, P=0.004; small variant, P=0.003). We also found an association of borderline significance (P=0.05) between the expression of RBM3, the large variant of RBM10 and wild-type p53. Expression of the small RBM10 variant, finally, was associated with high proliferation of the tumors (Ki67>or=20%; P=0.037). The expression of both RBM10 variants seems to be interdependent to a significant degree (r=0.26, P=0.006). From these results, it seems that the X-chromosome, through its RBM genes, plays a formerly unknown role in the regulation of programmed cell death (apoptosis) in breast cancer.

Characterization of a human Bid homologue protein from Gallus gallus.

Bid protein, a member of the "BH3-only" subgroup of Bcl-2 family, plays a critical role in mammalian apoptosis regulation. In this study, we have cloned the chicken Bid gene, which encodes a 193 amino acid protein and shares 40% homology with human and mouse Bid proteins. Bid sequence comparison emphasises the conservation of both the functional domain BH3 and the proteolytic cleavage sites. An induction of apoptosis by chicken Bid and the cleavage of the protein, after TNFalpha treatment, were also demonstrated. In addition, mRNA Bid expression was detected along all embryo stages and tissues examined, suggesting a role for this protein in the developmental process. This is the first report demonstrating the functionality of a "BH3-only" protein in chicken.

Proteomic analysis of the Gallus gallus embryo at stage-29 of development.

The chicken (Gallus gallus) is one of the primary models for embryological and developmental studies. In order to begin to understand the molecular mechanisms underlying the normal and abnormal development of the chicken, we used 2-DE to construct a whole-embryo proteome map. Proteins were separated by IEF on IPG strips, and by 11% SDS-PAGE) gels. Protein identification was performed by means of PMF with MALDI-TOF-MS. In all, 105 protein spots were identified, 35 of them implicated in embryo development, 10 related with some diseases, and 16, finally, being proteins that have never been identified, purified or characterized in the chicken before. This map will be updated continuously and will serve as a reference database for investigators, studying changes at the protein level under different physiological conditions.

CRR1, a gene encoding a putative transglycosidase, is required for proper spore wall assembly in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, sporulation is a developmental process that converts a single cell into four haploid spores. The four haploid nuclei are encapsulated within multilayered spore walls that protect them against stressful conditions. The formation of the spore-specific cell wall is a highly coordinated process that requires the participation of enzymic activities for biosynthesis, degradation, and cross-linking between components. Here the sporulation-specific gene CRR1, encoding a putative transglycosidase that is required for proper spore wall assembly, is described. Both the transcription of CRR1 and the synthesis of Crr1p were induced biphasically under sporulating conditions, with a first expression peak displaying kinetics similar to those of the middle to middle-late sporulation-specific genes, and a second late peak after 24 h under these conditions. Localization studies revealed that Crr1p localized to the spore wall that surrounds each of the four ascospores within the mature asci. Mutation of this gene had no effect on the efficiency of spore formation. However, crr1 mutant spores were sensitive to hydrolytic enzymes such as glusulase and to heat-shock treatments, underscoring the importance of this gene in the proper formation and assembly of the ascospore wall. Moreover, the deletion of CRR1 had additive effects with respect to the sensitivity of cda1 cda2 mutants to these treatments. Interestingly, overexpression of CRR1 not only complemented the phenotype of the crr1 strain but also rendered spores more resistant to the stress conditions than the wild-type. Like other mutants impaired in the formation of the spore outer layer, crr1 mutants were permeable to Calcofluor White. Finally, detailed analysis by electron microscopy of the spore walls in the crr1 mutants revealed a defect in the assembly of the spore wall components, suggesting a role for Crr1p in the cross-linking between the inner (glucan/mannoprotein) and the outer (chitosan/dityrosine) spore layers.

mRNA expression of the angiogenesis markers VEGF and CD105 (endoglin) in human breast cancer.

BACKGROUND: Both VEGF and CD105 (endoglin) have been identified as markers of tumor angiogenesis and prognosis in breast cancer. They have always been studied in this kind of tumor by means of immunological methods. OBJECTIVE: To study, by means of reverse-transcription polymerase chain reaction (RT-PCR), the expression of VEGF and CD105 (endoglin) at the mRNA level in a series of breast cancers and to correlate the results obtained with all available clinical and biological features of the tumors. MATERIALS AND METHODS: Fresh tumor tissue from 103 previously untreated breast cancer patients was studied for VEGF and CD105 (endoglin) expression. In addition, the following parameters were determined in all tumors: DNA ploidy by means of flow cytometry; hormone receptor (ER & PR), Ki67, c-erb-B2 and p53 expression by means of immunohistochemistry; and h-MAM (mammaglobin) expression by means of RT-PCR. Classical prognostic parameters of the tumors, such as histological and nuclear grade or axillary lymph node invasion, were also included into the statistical analysis. RESULTS: VEGF mRNA expression levels above the 25th percentile were significantly (p<0.05) associated with high proliferation (Ki67>10%) and aneuploidy of the tumors and inversely with estrogen receptor expression (p<0.01). CD105 (endoglin) mRNA expression levels above the 25th percentile only correlated significantly with nuclear grade 3 (p<0.05). The expression of both genes did not correlate with each other. CONCLUSION: VEGF mRNA expression levels seem to be directly associated with VEGF functions at the protein level, whereas this seems not to be the case for CD105 (endoglin) mRNA levels.

Nup88 mRNA overexpression is associated with high aggressiveness of breast cancer.

The nuclear pore complex protein Nup88 is overexpressed in tumor cells. Immunohistochemical studies have shown that this overexpression is linked to higher aggressiveness of colorectal carcinoma and to enhanced metastatic potential of melanoma cells. However, the antibodies so far developed against Nup88 have the drawback of recognizing a number of other, up to now unspecified antigens besides Nup88. For this reason, we devised the present study on Nup88 expression at the mRNA level. RNA was extracted from fresh tumor tissue corresponding to 122 breast cancer patients. Nup88 mRNA expression was measured by means of differential RT-PCR, standardizing against a constitutive internal control gene (beta-actin). The results were dichotomized into "high" and "low" expression levels, using the median value as cut-off. High Nup88 mRNA expression levels correlated significantly with ductal and tubular histology (p = 0.012), histologic and nuclear grade 3 of tumors (p < 0.001), absence of hormone receptor expression (p < 0.001), expression of the c-erb-B2 oncogene (p < 0.001), expression of mutant p53 protein (p < 0.001), high proliferation (defined by Ki67 labeling index >20%, p < 0.001), DNA aneuploidy (p < 0.001) as well as the most important ominous clinical prognostic factor, axillary node invasion (p < 0.001). We also found an inverse correlation (p < 0.001) with expression of the H-MAM (mammaglobin) gene, a marker of low biologic and clinical aggressiveness of breast cancer. All of these factors, without exception, define a highly aggressive tumor phenotype. These findings appear to be specific to Nup88 and not to nuclear pore proteins in general. Indeed, analysis of Nup107 (which is a limiting component of the nuclear pore complex) under the same conditions in the same tumors did not yield comparable results.