The association between duration of remission, fatigue, depression and health-related quality of life in Italian patients with systemic lupus erythematosus.

OBJECTIVE: To evaluate the impact of duration of remission on the health-related quality of life (HRQoL) of patients with systemic lupus erythematosus (SLE). METHODS: We conducted a 5-year retrospective study on two Italian cohorts. Remission was defined as a continuative period of no clinical disease activity, according to the Systemic Lupus Erythematosus Disease Activity Index 2 K, and a permitted maximum prednisone dose of 5 mg/day. HRQoL was measured using the 36-Item Short-Form Health Survey (SF36) during the last visit. RESULTS: We enrolled 136 female SLE patients. During observation, 15 (11%) patients had been in remission for ≥1 and <2 years, 15 (11%) for ≥2 and <3 years, 19 (14%) for ≥3 and <4 years, 9 (7%) for ≥4 and <5 years, and 53 (39%) had been in prolonged remission for ≥5 years. In the multivariate model, considering depression and fatigue as covariates, patients in prolonged remission showed significantly better scores in the physical functioning (p = 0.039), role physical (p = 0.029), bodily pain (p = 0.0057), general health (p = 0.0033) and social functioning (p = 0.0085) components of the SF36, compared with those in remission <5 years or unremitted. Subsequent mediation analyses found that these effects were partly influenced by depression. CONCLUSION: Lupus remission could improve the HRQoL of SLE patients, particularly when associated with appropriate management of depression and fatigue.

Health status and concomitant prescription of immunosuppressants are risk factors for hydroxychloroquine non-adherence in systemic lupus patients with prolonged inactive disease.

Background/objective The objectives of this paper are to assess the extent of and the factors associated with hydroxychloroquine (HCQ) non-adherence in systemic lupus erythematosus (SLE) patients with prolonged inactive disease and to investigate relationships between blood HCQ concentration and quality of life (QoL). Methods Consecutive SLE patients, in remission for at least one year and taking a stable dose of HCQ were investigated. At study entry (T0) and six months later (T6) a blood venous sample was taken to measure whole blood concentration of [HCQ] and desethylchloroquine ([DCQ]). Moreover, at T0 each patient completed validated questionnaires assessing QoL, disability, anxiety, depression and visual analogue scales for fatigue, pain, general health (GH), and self-assessment of disease activity. Results Eighty-three patients with a median [HCQ] of 327 ng/ml were enrolled. At T0, 24 (29%) were defined as non-adherent ([HCQ] < 100 ng/ml). At multiple logistic regression analysis the physical summary of SF-36 ( p = 0.038), and the concomitant use of immunosuppressants ( p = 0.010) were independently associated with non-adherence. A significant increase of HCQ adherence was observed at T6 ( p < 0.05). Conclusions A better health status and the concomitant prescription of immunosuppressants represent risk factors for HCQ non-adherence in SLE patients in remission. Monitoring HCQ levels might represent an important opportunity to improve adherence.

Primary prevention of cardiovascular disease in patients with systemic lupus erythematosus: case series and literature review.

Background Systemic lupus erythematosus is associated with an increased risk of cardiovascular disease. Low-dose aspirin, hydroxychloroquine and statins have been suggested to play a prophylactic role of cardiovascular events. This study is devoted to reviewing the literature on the topic and assessing the effects of these drugs in preventing a first cardiovascular event in a two-centre Italian series. Methods A PubMed search on cardiovascular prevention in systemic lupus erythematosus was performed. Moreover, systemic lupus erythematosus patients admitted to two centres from 2000-2015, who at admission had not experienced any cardiovascular event, were investigated. Aspirin, hydroxychloroquine and statin use, and the occurrence of any cardiovascular event, were recorded at each visit. Kaplan-Meier and Cox regression analyses were performed to evaluate the role of traditional, disease-related cardiovascular risk factors and of each of the three drugs in the occurrence of new cardiovascular events. Results The literature search produced conflicting results. Two hundred and ninety-one systemic lupus erythematosus patients were included in the study and followed for a median of eight years. During follow-up, 16 cardiovascular events occurred. At multivariate analysis, taking aspirin (hazard ratio: 0.24) and hydroxychloroquine for more than five years (hazard ratio: 0.27) reduced, while antiphospholipid antibody positivity (hazard ratio: 4.32) increased, the risk of a first cardiovascular event. No effect of statins emerged. Conclusion Our study confirms an additive role of aspirin and hydroxychloroquine in the primary prophylaxis of cardiovascular events in Italian patients with systemic lupus erythematosus. The lack of any detected effect in previous reports may depend on the design of studies and their short follow-up period.

Autoimmune gastrointestinal complications in patients with systemic lupus erythematosus: case series and literature review.

The association of systemic lupus erythematosus (SLE) with gastrointestinal autoimmune diseases is rare, but has been described in the literature, mostly as case reports. However, some of these diseases may be very severe, thus a correct and early diagnosis with appropriate management are fundamental. We have analysed our data from the SLE patient cohort at University College Hospital London, established in 1978, identifying those patients with an associated autoimmune gastrointestinal disease. We have also undertaken a review of the literature describing the major autoimmune gastrointestinal pathologies which may be coincident with SLE, focusing on the incidence, clinical and laboratory (particularly antibody) findings, common aetiopathogenesis and complications.

Expression and localization of the deubiquitinating enzyme mUBPy in wobbler mouse testis during spermiogenesis.

Mouse ubiquitin-specific processing protease (mUBPy) is a deubiquitinating enzyme highly expressed in both brain and testis. In testis, it interacts with the DnaJ protein, MSJ-1; both mUBPy and MSJ-1 are located on the cytoplasmic surface of the developing acrosome and in the centrosomal region during spemiogenesis. Present data show the first appearance in testis of mUbpy mRNA and protein at 10 days post-partum (d.p.p.). In addition, to investigate on a possible role of mUBPy in sperm formation, we took advantage of mutant wr/wr (wobbler) mice characterized by male infertility, which is likely due to the lack of a real, functional acrosome. RT-PCR and Northern blot analyses show that mUbpy is up-regulated in adult wobbler testis. Furthermore, in wild-type testis mUBPy protein is primarily detected by Western blot in the soluble (cytosolic/nuclear) fraction during the first round of spermatogenesis and in the adult. By contrast, mUBPy is primarily detected in membranous/insoluble protein fraction when wobbler phenotype is clearly shown (30 d.p.p.) and in adult wobbler testis. By immunohistochemistry, whereas in wild-type animals mUBPy marks the profile of the acrosomic vesicle in differentiating spermatids, in wobbler mice only a detergent pre-treatment procedure allows to detect mUBPy immunoreactivity, which results in diffuse spotted granules inside the cytoplasm and around the nuclear shape. In conclusion, in wobbler testis expression of mUbpy is up-regulated, while a differential sorting of the protein characterizes wobbler spermatids where acrosome formation is impaired.

Multipotential neural precursors transplanted into the metachromatic leukodystrophy brain fail to generate oligodendrocytes but contribute to limit brain dysfunction.

Neural stem cells appear to be best suited for regenerative therapy in neurological diseases. However, the effects of high levels of potentially toxic substances such as sulfatides--which accumulate in metachromatic leukodystrophy (MLD)--on this regenerative ability are still largely unclear. To start addressing this question, in vitro and in vivo experiments were used to examine the behavior of multipotential neural precursors exposed to abnormally high levels of sulfatides. Following transplantation of dissociated neurospheres into the brain of presymptomatic MLD pups, the majority of donor-derived cells were distributed in a caudal to rostral direction, with higher numbers in the cortex. Most if not all of the donor cells acquired an astroglial phenotype. We found no evidence of oligodendrocyte or neuronal commitment of transplanted cells in long-term-treated MLD mice (e.g. up to 1.5 years of age). This was in line with our in vitro findings of sulfatides blocking oligodendrocyte formation after induction of differentiation in sulfatide-treated epidermal growth factor/fibroblast growth factor responsive neurospheres. Transplanted MLD mice showed an improved arylsulfatase A (ARSA) activity and a significant amelioration of sulfatide metabolism, neurodegeneration and motor-learning/memory deficits. Furthermore, transplanted cells were shown to act as a source of ARSA enzyme that accumulated in endogenous brain cells, indicating the occurrence of enzyme cross-correction between transplanted and host cells. These results provide a first insight into the effect of sulfatides on the stemness properties of neural stem cells and on the effects of the MLD environment on the in vivo expectations of using neural stem cells in cell therapy.

Characterization of Follicular Atresia Responsive to BPA in Zebrafish by Morphometric Analysis of Follicular Stage Progression.

Bisphenol A is an industrial chemical compound, pervasively polluting the environment and diet, classified as an endocrine disruptor because of its interference effects on the endocrine system. In zebrafish, BPA exposure induces follicular atresia. To acquire knowledge on this atretic effect, using a qualitative and quantitative histomorphological approach, we studied zebrafish ovarian follicular stage development in response to low BPA concentrations. Results show that BPA interferes with follicular progression by affecting the previtellogenic and vitellogenic phases. In particular, BPA exposure (i) increases follicular recruitment by acting on primary stage follicles, (ii) forces the follicular transition from stage III to stage IV producing enlarged stage IV follicles, and (iii) induces atresia by producing atretic follicles that are peculiarly enlarged (i.e., big atretic follicles). We suggest that BPA induces atresia by the primary effect on recruitment of stage I follicles. This forces follicular progression and produces stage IV follicles that are peculiarly enlarged that undertake the atretic development.

Safety of arylsulfatase A overexpression for gene therapy of metachromatic leukodystrophy.

Successful gene therapy approaches for metachromatic leukodystrophy (MLD), based either on hematopoietic stem/progenitor cells (HSPCs) or direct central nervous system (CNS) gene transfer, highlighted a requirement for high levels of arylsulfatase A (ARSA) expression to achieve correction of disease manifestations in the mouse model. Full assessment of the safety of ARSA expression above physiological levels thus represents a prerequisite for clinical translation of these approaches. Here, using lentiviral vectors (LVs), we generated two relevant models for the stringent evaluation of the consequences of ARSA overexpression in transduced cells. We first demonstrated that ARSA overexpression in human HSPCs does not affect their clonogenic and multilineage differentiation capacities in clonogenic assays and in a neonatal hematochimeric mouse model. Further, we studied ARSA overexpression in all body tissues by generating transgenic mice overexpressing the ARSA enzyme by LV up to 15-fold above the normal range and carrying multiple copies of LV in their genome. Characterization of these mice demonstrated the safety of ARSA overexpression in two main gene therapy targets, HSPCs and neurons, with maintenance of the complex functions of the hematopoietic and nervous system in the presence of supraphysiological enzyme levels. The activity of other sulfatases dependent on the same common activator, sulfatase-modifying factor-1 (SUMF1), was tested in ARSA-overexpressing HSPCs and in transgenic mice, excluding the occurrence of saturation phenomena. Overall, these data indicate that from the perspective of clinical translation, therapeutic levels of ARSA overexpression can be safely achieved. Further, they demonstrate an experimental platform for the preclinical assessment of the safety of new gene therapy approaches.

Structure of msj-1 gene: a comparative analysis.

Msj-1 gene encodes a DnaJ protein highly expressed in spermatids and spermatozoa of both rodents and amphibians. We isolated and characterized the msj-1 gene in mice. A bioinformatic approach was then used to predict the putative promoter region, chromosomal localization, and its presence in the human genome. The analysis of msj-1 genomic sequence revealed that msj-1 is an intronless gene. Interestingly, two regions (A and B, separated by 10,682 bp) on human chromosome 2 having respectively 78% and 77% nucleotide identity with the murine msj-1 coding region were identified. This suggests the existence of an msj-1-like gene also in humans.

Cellular mechanisms of striatum-dependent behavioral plasticity and drug addiction.

The striatum has long been known to be involved in the control of motor behavior, since disruption of dopamine-mediated function in this brain structure is directly linked to Parkinson's disease and other disorders of movement. However, it is now accepted that both dorsal and ventral striatal nuclei are also essential for a variety of cognitive processes, which depend on reward-based stimulus-response learning. Since the neuroanatomical and neurochemical organization of dorsal and ventral striatum is only partially overlapping, it is likely that both common and nucleus-specific cellular and molecular events contribute to synaptic plasticity, learning and memory processes mediated by these cerebral structures. Alterations in cell signaling in the striatum may be particularly important in the response to both acute and chronic administration of drugs of abuse, resulting in maladaptive changes in the reward-based associative learning involved in addiction, withdrawal and relapse.

A gonadotropin-releasing hormone (GnRH) antagonist decreases androgen production and spermatogonial multiplication in frog (Rana esculenta): indirect evidence for the existence of GnRH or GnRH-like material receptors in the hypophysis and testis.

The effects of a GnRH antagonist (GnRHA) on GnRH agonist (GnRH*)-induced androgen production and spermatogonial multiplication were studied in the frog, Rana esculenta, in vivo and in vitro. Intact and hypophysectomized (PDX) animals were kept at 22 +/- 2 C and treated with GnRH (45 ng/g BW) and GnRH* plus 1X and 10X concentrations of GnRHA on alternate days for 2 weeks. Androgen concentration in GnRH* plus GnRHA-treated animals decreased in the testis by about 50% with the 10X dose whereas the increase obtained in GnRH*-treated PDX group was completely abolished with the 1X dose. Histological sections were evaluated with respect of the mitotic index (MI) of the primary spermatogonia. Both GnRHA-treated intact and PDX frogs showed a dose-dependent MI decrease which reached 59% and 57% of control, respectively. In vitro incubations were carried out on testis halves at 15 C for 0, 2, 4, 6, and 8 h with the addition of 1 microgram GnRH* and 1 microgram GnRH* plus 1 or 10 micrograms GnRHA. The stimulatory effect of GnRH* and the inhibitory effect of GnRHA were apparent within 2 h. The basal mitogenic activity was affected by antagonist treatment and the inhibitory effect on the MI was evident within 2-4 h in the 10X-treated groups or within 6-8 h in the 1X treated groups. Since GnRH* and GnRHA bind to the same receptor these data strongly indicate that the effects of putative GnRH-like materials in the frog, Rana esculenta, are mediated throughout stereospecific recognition sites in both pituitary and testis.

c-fos activity in Rana esculenta testis: seasonal and estradiol-induced changes.

Estradiol-17beta (E2) is suspected to exert a role in the regulation of testicular activity. Using a nonmammalian vertebrate model (the frog, Rana esculenta), we have investigated whether c-fos activity is detectable in the testis during the annual sexual cycle and whether E2 exerts a regulatory role on spermatogenesis through fos activity. FOS protein is available in testicular nuclear extracts (about 60 kDa) and, surprisingly, also in cytosolic extracts (about 60, 80, and 100 kDa). Estradiol induces primary spermatogonia (ISPG) proliferation [this effect is counteracted by antiestrogens (Tamoxifen and ICI 182-780)] and FOS appearance in testicular cytosolic extracts as well as c-fos transcription. Also, this effect is counteracted by ICI 182-780. Interestingly, the number of FOS immunopositive nuclei of ISPG strongly increases after E2 treatment, whereas a great increase of immunopositivity in the cytoplasm of ISPG is observed with the contemporaneous treatment with antiestrogens. In conclusion, our results demonstrate that E2 induces ISPG multiplication in the frog, R. esculenta, and, for the first time in a vertebrate species, that it triggers c-fos activity in the testis. Moreover, E2 may be involved in mechanisms related to FOS transport in the nucleus of ISPG to induce the mitotic activity.

Early defect in the expression of mouse sperm DNAJ 1, a member of the DNAJ/heat shock protein 40 chaperone protein family, in the spinal cord of the wobbler mouse, a murine model of motoneuronal degeneration.

Prevention of protein misfolding is ensured by chaperone proteins, including the heat shock proteins (HSP) of the DNAJ/HSP40 family. Detection of abnormal protein aggregates in various neurodegenerative diseases has led to the proposal that altered chaperone activity contributes to neurodegeneration. Msj-1, a DNAJ/HSP40 protein located around the spermatozoa acrosome, was recently found to be down-regulated in the testis of wobbler mutant mice. Wobbler is an unidentified recessive mutation which triggers progressive motoneuron degeneration with abnormal intracellular protein accumulations, and defective spermatozoa maturation. Here, we examined Msj-1 expression in the spinal cord of the mutants and their controls. Msj-1 transcripts were amplified by reverse transcription-polymerase chain reaction from mutant and wild-type spinal cord RNA. Sequencing of Msj-1 coding region revealed no change in the mutant. In contrast, decreased Msj-1 mRNA levels were observed in five to six-week-old wobbler mice spinal cord, when motoneuron degeneration is at its apex, as compared to controls. A similar decrease was observed in two-week-old wobbler spinal cord, when the number of motoneurons is still unaltered, indicating that the decreased mRNA content is intrinsic to the mutant and not simply related to the loss of cells expressing Msj-1. Assays of Msj-1 protein levels yielded similar results. Immunofluorescent labeling revealed numerous Msj-1-ir motoneurons in five-week-old control spinal cord while no signal was observed in age-matched wobbler. Our results show, therefore, that Msj-1 expression is down-regulated in both organs affected by the wobbler mutation, the CNS and the testis, and that this defect precedes the first histological signs of motoneuron degeneration. These results provide the first example of an association between transcriptional repression of a chaperone protein and a neurodegenerative process.

Seasonal testosterone profile and testicular responsiveness to pituitary factors and gonadotrophin releasing hormone during two different phases of the sexual cycle of the frog (Rana esculenta).

Plasma and testicular testosterone concentrations in the frog, Rana esculenta, were studied by radioimmunoassay and showed similar seasonal fluctuations. The increase in testicular androgen during November preceded that occurring in the plasma by 2 months. Pituitary products and gonadotrophin releasing hormone, and the responsiveness of the testis to these substances play an important role in determining the hormone profile.

Opioids and testicular activity in the frog, Rana esculenta.

The presence and activity of brain, pituitary and testicular beta-endorphin (beta-EP)-like material have been studied in the frog, Rana esculenta, using reverse-phase high-pressure liquid chromatography, coupled with radioimmunoassay and immunocytochemistry. In-vivo and in-vitro treatments with naltrexone were carried out to assess the putative physiological activity of opioid peptides. beta-EP(1-31) and (1-27), together with their acetylated forms, have been identified in brain, pituitary and testis. In particular, beta-EP(1-31) concentrations peaked during July in the brain and pituitary, whilst in testes maximum concentrations were found in April and November. beta-EP immunoreactivity was present in the brain within the nucleus preopticus and nucleus infundibularis ventralis while positive fibres in the retrochiasmatic regions projected to the median eminence. In the testis, interstitial cells, canaliculi of the efferent system, spermatogonia and spermatocytes showed positive immunostaining for beta-EP. In intact animals, naltrexone treatment increased plasma and testicular androgen levels and this effect was confirmed in in-vitro incubations of minced testes. Naltrexone also induced a significant increase in germ cell degeneration. Our results indicated that an opioid system modulates the hypothalamus-pituitary-gonadal axis in the frog, Rana esculenta and, for the first time, we have shown that the testicular activity of a non-mammalian species may be regulated by opiates locally.

Opioid peptides and testicular activity in the lizard Podarcis s. sicula Raf.

In mammals endorphinergic systems have been shown to modulate reproductive processes and beta-endorphin (beta-EP) has been found to influence sexual functions, acting at the hypothalamus-pituitary-gonadal axis level. Using immunocytochemical and in vitro studies, evidence for a diffuse pro-opiomelanocortin-related opioid system in the lizard Podarcis s. sicula was produced. In the testis, beta-EP immunoreactivity showed seasonal variation, being most pronounced in the interstitial cells of sexually quiescent lizards (December). Reverse-phase high-performance liquid chromatography, coupled with radioimmunoassay and immunocytochemistry, showed that beta-EP and acetyl beta-EP increased during December, while their concentrations were low during April, when the highest testicular activity occurred. Using in vivo studies, it was found that naltrexone treatment, blocking pituitary opioid receptor, increased androgen levels in the plasma and in the testis. It was also found with in vitro studies that the endogenous opioid system inhibits gonadotrophin release and therefore androgen production by the testis. The data reported here provide evidence for the physiological role played by opioid peptides at the pituitary level to regulate the seasonal reproductive activity of the lizard Podarcis s. sicula.

Detection of c-mos related products in the dogfish (Scyliorhinus canicula) testis.

The objective of the present paper was to do a comparative study to assess somatic versus germ cell localization of c-mos products in the testis. In mouse and amphibian oocytes, c-mos activity is necessary for meiotic maturation. Lack of c-mos expression has been reported in somatic cells of male and female gonads while transcripts have been found in germ cells of testis and ovary. Using a v-mos probe, we report here the detection of a c-mos related transcript (1.7 kb) in the dogfish Scyliorhinus canicula testis. Western blot analysis detects two proteins of 106 and 32 kDa. A specific immunostaining was exclusively localized in the interstitial tissue while the germinal compartment was completely negative. In conclusion, our results indicate for the first time the presence of c-mos products in an elasmobranch species and, moreover, their presence in somatic testicular cells rather than germ cells. Therefore, this finding in an ancient vertebrate indicates that c-mos activity does not have a direct universal role in the regulation of spermatogenesis.

Stimulatory effect of a GnRH agonist (buserelin) in in vitro and in vivo testosterone production by the frog (Rana esculenta) testis.

The summary testicular effects of an agonistic analogue of gonadotropin-releasing hormone (buserelin, GnRHa) have been studied in vitro and in vivo in the frog, Rana esculenta. During 3 h incubation GnRHa (8 X 10(-7) M) potentiated pituitary factors in stimulating testosterone production by minced testes in vitro. After 6 h of incubation 8 X 10(-7) M GnRHa stimulated maximal testosterone output. Testes of 10-day hypophysectomized animals did not show any GnRHa effect in vitro. In vivo, a direct effect of GnRHa on testicular testosterone production was demonstrated in hypophysectomized animals, although this effect was temperature-dependent, requiring the frog to be maintained at a high temperature (24 degrees C). No effect of GnRHa was detectable in frogs kept at a low temperature (4 degrees C).

Fos localization in cytosolic and nuclear compartments in neurones of the frog, Rana esculenta, brain: an analysis carried out in parallel with GnRH molecular forms.

C-fos activity was determined in the brain of the frog, Rana esculenta, during the annual sexual cycle. The localization of GnRH molecular forms (mammalian- and chicken-GnRHII) was also carried out to determine whether or not the proto-oncogene and the peptides showed a functional relationship. Northern blot analysis of total RNA revealed the presence of a single strong signal of c-fos like mRNA of 1.9 Kb during February and April. This was followed by expression of c-Fos protein (Fos) in several brain areas during March and July shown by immunocytochemistry. In particular, the olfactory region, the lateral and medial pallium, the nucleus lateralis septi, the ventral striatum, the caudal region of the anterior preoptic area, the suprachiasmatic nucleus, the ventral thalamus, tori semicircularis and ependymal layers of the tectum were immunostained. There was no overlap between Fos immunoreactive perikarya and GnRH immunoreactive perikarya (e.g. gonadotrophin-releasing hormone (GnRH) in the rostral part and Fos in the caudal region of the anterior preoptic area). Interestingly, a cytoplasmic localization of Fos was also observed by immunocytochemistry and gel retardation experiments supported this observation. Cytoplasmic extracts from September-October animals bound the AP1 oligonucleotide. The complex was not available in the nuclear extracts from the same preparation, suggesting that, besides Fos, Jun products were also present. Conversely, nuclear but not cytosolic binding was detected in the brain of animals collected in July. In conclusion, we show that Fos and GnRH activity does not correlate in the frog brain and, for the first time in a vertebrate species, we give evidence of a cytoplasmic AP1 complex in neuronal cells.