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Introduction: The pathogenesis of neuropsychiatric systemic lupus erythematosus (NPSLE) is widely unknown, and the role of autoantibodies is still undetermined. Methods: To identify brain-reactive autoantibodies possibly related to NPSLE, immunofluorescence (IF) and transmission electron microscopy (TEM) on rat and human brains were performed. ELISA was used to reveal the presence of known circulating autoantibodies, while western blot (WB) was applied to characterize potential unknown autoantigen(s). Results: We enrolled 209 subjects, including patients affected by SLE (n=69), NPSLE (n=36), Multiple Sclerosis (MS, n=22), and 82 age- and gender-matched healthy donors (HD). Autoantibody reactivity by IF was observed in almost the entire rat brain (cortex, hippocampus, and cerebellum) using sera from NPSLE and SLE patients and was virtually negative in MS and HD. NPSLE showed higher prevalence (OR 2.4; p = 0.047), intensity, and titer of brain-reactive autoantibodies than SLE patients. Most of the patient sera with brain-reactive autoantibodies (75%) also stained human brains. Double staining experiments on rat brains mixing patients' sera with antibodies directed against neuronal (NeuN) or glial markers showed autoantibody reactivity restricted to NeuN-containing neurons. Using TEM, the targets of brain-reactive autoantibodies were located in the nuclei and, to a lesser extent, in the cytoplasm and mitochondria. Given the high degree of colocalization between NeuN and brain-reactive autoantibodies, we assumed NeuN was a possible autoantigen. However, WB analysis with HEK293T cell lysates expressing or not expressing the gene encoding for NeuN protein (RIBFOX3) showed that patients' sera carrying brain-reactive autoantibodies did not recognize the NeuN corresponding band size. Among the panel of NPSLE-associated autoantibodies (e.g., anti-NR2, anti-P-ribosomal protein, antiphospholipid) investigated by ELISA assay, only the anti-b2- glycoprotein-I (ab2GPI) IgG was exclusively found in those sera containing brain-reactive autoantibodies. Conclusion: In conclusion, SLE and NPSLE patients possess brain-reactive autoantibodies but with higher frequency and titers found in NPSLE patients. Although many target antigens of brain-reactive autoantibodies are still undetermined, they likely include β2GPI. [ABSTRACT FROM AUTHOR]

Colorectal cancer (CRC) is the fourth cause of death from cancer worldwide mainly due to the high incidence of drug‐resistance. During a screen for new actionable targets in drug‐resistant tumours we recently identified p65BTK – a novel oncogenic isoform of Bruton's tyrosine kinase. Studying three different cohorts of patients here we show that p65BTK expression correlates with histotype and cancer progression. Using drug‐resistant TP53‐null colon cancer cells as a model we demonstrated that p65BTK silencing or chemical inhibition overcame the 5‐fluorouracil resistance of CRC cell lines and patient‐derived organoids and significantly reduced the growth of xenografted tumours. Mechanistically, we show that blocking p65BTK in drug‐resistant cells abolished a 5‐FU‐elicited TGFB1 protective response and triggered E2F‐dependent apoptosis. Taken together, our data demonstrated that targeting p65BTK restores the apoptotic response to chemotherapy of drug‐resistant CRCs and gives a proof‐of‐concept for suggesting the use of BTK inhibitors in combination with 5‐FU as a novel therapeutic approach in CRC patients. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

TLQP-21, a peptide encoded by the highly conserved vgf gene, is expressed in neuroendocrine cells and has been the most prominent VGF-derived peptide studied in relation to control of energy balance. The recent discovery that TLQP-21 is the natural agonist for the complement 3a receptor 1 (C3aR1) has revived interest in this peptide as a potential drug target for obesity. We have investigated its function in Siberian hamsters (Phodopus sungorus), a rodent that displays natural seasonal changes in body weight and adiposity as an adaptation to survive winter. We have previously shown that intracerebroventricular administration of TLQP-21 reduced food intake and body weight in hamsters in their long-day fat state. The aim of our current study was to determine the systemic actions of TLQP-21 on food intake, energy expenditure and body weight, and to establish whether adiposity affected these responses. Peripheral infusion of TLQP-21 (1mg/kg/day for 7 days) in lean hamsters exposed to short photoperiods (SP) reduced cumulative food intake in the home cage (p<0.05), and intake when measured in metabolic cages (P<0.01). Energy expenditure was significantly increased (p<0.001) by TLQP-21 infusion, this was associated with a significant increase in uncoupling protein 1 mRNA in brown adipose tissue (BAT) (p<0.05), and body weight was significantly reduced (p<0.05). These effects of systemic TLQP-21 treatment were not observed in hamsters exposed to long photoperiod (LP) with a fat phenotype. C3aR1 mRNA and protein were abundantly expressed in the hypothalamus, brown and white adipose tissue in hamsters, but changes in expression cannot explain the differential response to TLQP-21 in lean and fat hamsters. [ABSTRACT FROM AUTHOR]

This review summarized different studies reporting the presence of autoantibodies reacting against cells of the pituitary (APAs) and/or hypothalamus (AHAs). Both APAs and AHAs have been revealed through immunofluorescence using different kinds of substrates. Autoantibodies against gonadotropic cells were mainly found in patients affected by cryptorchidism and hypogonadotropic hypogonadism while those against prolactin cells were found in different kinds of patients, the majority without pituitary abnormalities. APAs to growth hormone (GH) cells have been associated with GH deficiency while those against the adrenocorticotropic cells have distinguished central Cushing's disease patients at risk of incomplete cure after surgical adenoma removal. AHAs to vasopressin cells have identified patients at risk of developing diabetes insipidus. APAs have been also found together with AHAs in patients affected by idiopathic hypopituitarism, but both were also present in different kinds of patients without abnormalities of the hypothalamic-pituitary axis. Despite some data being promising, the clinical use of pituitary and hypothalamus autoantibodies is still limited by the low diagnostic sensitivity, irreproducibility of the results, and the absence of autoantigen/s able to discriminate the autoimmune reaction involving the pituitary or the hypothalamus from the other autoimmune states. [ABSTRACT FROM AUTHOR]

From the VGF precursor protein originate several low molecular weight peptides, whose distribution in the brain and blood circulation is not entirely known. Among the VGF peptides, those containing the N-terminus portion were altered in the cerebro-spinal fluid (CSF) and hypothalamus of schizophrenia patients. "Hence, we aimed to better investigate the involvement of the VGF peptides in schizophrenia by studying their localization in the brain regions relevant for the disease, and revealing their possible modulations in response to certain neuronal alterations occurring in schizophrenia". We produced antibodies against different VGF peptides encompassing the N-terminus, but also C-terminus-, TLQP-, GGGE- peptide sequences, and the so named NERP-3 and-4. These antibodies were used to carry out specific ELISA and immunolocalization studies while mass spectrometry (MS) analysis was also performed to recognize the intact brain VGF fragments. We used a schizophrenia rat model, in which alterations in the prepulse inhibition (PPI) of the acoustic startle response occurred after PCP treatment. In normal rats, all the VGF peptides studied were distributed in the brain areas examined including hypothalamus, prefrontal cortex, hippocampus, accumbens and amygdaloid nuclei and also in the plasma. By liquid chromatography-high resolution mass, we identified different intact VGF peptide fragments, including those encompassing the N-terminus and the NERPs. PCP treatment caused behavioral changes that closely mimic schizophrenia, estimated by us as a disruption of PPI of the acoustic startle response. The PCP treatment also induced selective changes in the VGF peptide levels within certain brain areas. Indeed, an increase in VGF C-terminus and TLQP peptides was revealed in the prefrontal cortex (p < 0.01) where they were localized within parvoalbumin and tyrosine hydroxylase (TH) containing neurons, respectively. Conversely, in the nucleus accumbens, PCP treatment produced a down-regulation in the levels of VGF C-terminus-, N-terminus- and GGGEpeptides (p < 0.01), expressed in GABAergic- (C-terminus/GGGE) and somatostatin- (N-terminus) neurons. These results confirm that VGF peptides are widely distributed in the brain and modulated in specific areas involved in schizophrenia. [ABSTRACT FROM AUTHOR]

VGF (non-acronymic) was first highlighted to have a role in energy homeostasis through experiments involving dietary manipulation in mice. Fasting increased VGF mRNA in the Arc and levels were subsequently reduced upon refeeding. This anabolic role for VGF was supported by observations in a VGF null (VGF-/-) mouse and in the diet-induced and gold-thioglucose obese mice. However, this anabolic role for VGF has not been supported by a number of subsequent studies investigating the physiological effects of VGF-derived peptides. Intracerebroventricular (ICV) infusion of TLQP-21 increased resting energy expenditure and rectal temperature in mice and protected against diet-induced obesity. Similarly, ICV infusion of TLQP-21 into Siberian hamsters significantly reduced body weight, but this was due to a decrease in food intake, with no effect on energy expenditure. Subsequently NERP-2 was shown to increase food intake in rats via the orexin system, suggesting opposing roles for these VGF-derived peptides. Thus to further elucidate the role of hypothalamic VGF in the regulation of energy homeostasis we utilised a recombinant adeno-associated viral vector to over-express VGF in adult male Siberian hamsters, thus avoiding any developmental effects or associated functional compensation. Initially, hypothalamic over-expression of VGF in adult Siberian hamsters produced no effect on metabolic parameters, but by 12 weeks post-infusion hamsters had increased oxygen consumption and a tendency to increased carbon dioxide production; this attenuated body weight gain, reduced interscapular white adipose tissue and resulted in a compensatory increase in food intake. These observed changes in energy expenditure and food intake were associated with an increase in the hypothalamic contents of the VGF-derived peptides AQEE, TLQP and NERP-2. The complex phenotype of the VGF-/- mice is a likely consequence of global ablation of the gene and its derived peptides during development, as well as in the adult. [ABSTRACT FROM AUTHOR]

VGF mRNA is widely expressed in areas of the nervous system known to degenerate in Amyotrophic Lateral Sclerosis (ALS), including cerebral cortex, brainstem and spinal cord. Despite certain VGF alterations are reported in animal models, little information is available with respect to the ALS patients. We addressed VGF peptide changes in fibroblast cell cultures and in plasma obtained from ALS patients, in parallel with spinal cord and plasma samples from the G93A-SOD1 mouse model. Antisera specific for the C-terminal end of the human and mouse VGF proteins, respectively, were used in immunohistochemistry and enzyme-linked immunosorbent assay (ELISA), while gel chromatography and HPLC/ESI-MS/MS were used to identify the VGF peptides present. Immunoreactive VGF C-terminus peptides were reduced in both fibroblast and plasma samples from ALS patients in an advanced stage of the disease. In the G93A-SOD1 mice, the same VGF peptides were also decreased in plasma in the late-symptomatic stage, while showing an earlier down-regulation in the spinal cord. In immunohistochemistry, a large number of gray matter structures were VGF C-terminus immunoreactive in control mice (including nerve terminals, axons and a few perikarya identified as motoneurons), with a striking reduction already in the pre-symptomatic stage. Through gel chromatography and spectrometry analysis, we identified one form likely to be the VGF precursor as well as peptides containing the NAPP- sequence in all tissues studied, while in the mice and fibroblasts, we revealed also AQEE- and TLQP- peptides. Taken together, selective VGF fragment depletion may participate in disease onset and/or progression of ALS. [ABSTRACT FROM AUTHOR]

To address the possible involvement of VGF peptides in obesity and diabetes, we studied type 2 diabetes (T2D) and obese patients, and high-fat diet induced obese mice. Two VGF peptides (NAPP-19 and QQET-30) were identified in human plasma by HPLC-ESI-MS. The VGF C-terminus, the above two cleaved peptides, and the TLQP-21 related peptide/s were studied using ELISA and immunohistochemistry. In euglycemic patients, plasma NAPPE and TLQP like peptides were significantly reduced with obesity (74±10 vs. 167±28, and 92±10 vs. 191±19 pmol/ml, mean+SEM, n = 10 and 6, obese vs. normal BMI, respectively, p<0.03). Upon a standard glucose load, a distinct response was shown for VGF C-terminus, TLQP and QQET-like (ERVW immunoreactive) peptides in euglycemic normal BMI patients, but was virtually abolished in euglycemic obese, and in T2D patients independently of BMI. High-fat diet induced obese mice showed reduced plasma VGF C-terminus, NAPPE and QQET-like (ERVW) peptide/s (3±0.2 vs. 4.6±0.3, 22±3.5 vs. 34±1.3, and 48±7 vs. 100±7 pmol/ml, mean+SEM, n = 8/group, obese vs. slim, respectively, p<0.03), with a loss of the response to glucose for all VGF peptides studied. In immunohistochemistry, TLQP and/or VGF C-terminus antibodies labelled VGF containing perikarya in mouse celiac ganglia, pancreatic islet cells and thin beaded nerve fibres in brown adipose tissues, with fewer in white adipose tissue. Upon the glucose load, tyrosine hydroxylase and VGF C-terminus immunoreactive axons became apparent in pancreatic islets of slim animals, but not in obese animals. Alltogether, a significant loss of VGF peptide immunoreactivity and/or their response to glucose was demonstrated in obese patients, with or without T2D, in parallel with a similar loss in high-fat diet induced obese mice. An involvement of VGF in metabolic regulations, including those of brown and/or white adipose tissues is underlined, and may point out specific VGF peptides as potential targets for diagnosis and/or treatment. [ABSTRACT FROM AUTHOR]

VGF mRNA is induced in specific hypothalamic areas of the Siberian hamster upon exposure to short photoperiods, which is associated with a seasonal decrease in appetite and weight loss. Processing of VGF generates multiple bioactive peptides, so the objective of this study was to determine the profile of the VGF-derived peptides in the brain, pituitary and plasma from Siberian hamsters, and to establish whether differential processing might occur in the short day lean state versus long day fat. Antisera against short sequences at the C- or N- termini of proVGF, as well as against NERP-1, TPGH and TLQP peptides, were used for analyses of tissues, and both immunohistochemistry and enzyme linked immunosorbent assay (ELISA) coupled with high-performance liquid (HPLC) or gel chromatography were carried out. VGF peptide immunoreactivity was found within cortex cholinergic perikarya, in multiple hypothalamic nuclei, including those containing vasopressin, and in pituitary gonadotrophs. ELISA revealed that exposure to short day photoperiod led to a down-regulation of VGF immunoreactivity in the cortex, and a less pronounced decrease in the hypothalamus and pituitary, while the plasma VGF levels were not affected by the photoperiod. HPLC and gel chromatography both confirmed the presence of multiple VGF-derived peptides in these tissues, while gel chromatography showed the presence of the VGF precursor in all tissues tested except for the cortex. These observations are consistent with the view that VGF-derived peptides have pleiotropic actions related to changing photoperiod, possibly by regulating cholinergic systems in the cortex, vasopressin hypothalamic pathways, and the reproductive axis. [ABSTRACT FROM AUTHOR]

Although the VGF derived peptide TLQP-21 stimulates gonadotropin-releasing hormone (GnRH) and gonadotropin secretion, available data on VGF peptides and reproduction are limited. We used antibodies specific for the two ends of the VGF precursor, and for two VGF derived peptides namely TLQP and PGH, to be used in immunohistochemistry and enzyme-linked immunosorbent assay complemented with gel chromatography. In cycling female rats, VGF C-/N-terminus and PGH peptide antibodies selectively labelled neurones containing either GnRH, or kisspeptin (VGF N-terminus only), pituitary gonadotrophs and lactotrophs, or oocytes (PGH peptides only). Conversely, TLQP peptides were restricted to somatostatin neurones, gonadotrophs, and ovarian granulosa, interstitial and theca cells. TLQP levels were highest, especially in plasma and ovary, with several molecular forms shown in chromatography including one compatible with TLQP-21. Among the cycle phases, TLQP levels were higher during metestrus-diestrus in median eminence and pituitary, while increased in the ovary and decreased in plasma during proestrus. VGF N- and C-terminus peptides also showed modulations over the estrous cycle, in median eminence, pituitary and plasma, while PGH peptides did not. In ovariectomised rats, plasmatic TLQP peptide levels showed distinct reduction suggestive of a major origin from the ovary, while the estrogen-progesterone treatment modulated VGF C-terminus and TLQP peptides in the hypothalamus-pituitary complex. In in vitro hypothalamus, TLQP-21 stimulated release of growth hormone releasing hormone but not of somatostatin. In conclusion, various VGF peptides may regulate the hypothalamus-pituitary complex via specific neuroendocrine mechanisms while TLQP peptides may act at further, multiple levels via endocrine mechanisms involving the ovary. [ABSTRACT FROM AUTHOR]

Context. Although pituitary autoantibodies have frequently been reported in Autoimmune Polyendocrine Syndrome type 1 (APS1), the autoimmune involvement of the hypothalamic-pituitary axis remains to be elucidated. Objective. Our aim was to identify in APS1 patients novel autoantibodies, especially against hypothalamic-pituitary targets, and to correlate their presence with clinical features. Patients. We analyzed 14 APS1 patients from Sardinia, compared to other diseases and healthy donors. Measure(s). We used immunohistochemistry, on tissues substrates from various neuroendocrine organs, to detect autoantibody targets. Immunoenzymatic assays, as well as absorption with specific antigens were used to reveal autoantibodies against growth hormone (GH), luteinizing hormone (LH) and somatocrinin (GHRH). Clinical evaluations included GH secretory and cardiovascular autonomic neuropathy tests. Results. Sera from 12/14 APS1 patients revealed autoantibodies reacting with the hypothalamic-pituitary axis, cerebellum, substantia nigra, and/or adrenal medulla, as well as with GH, LH and/or GHRH. Of APS1 patients, 5 showed GH deficiency, in association (4/5 cases) with autoantibodies to hypothalamic and/or pituitary targets. Hypogonadotrophic hypogonadism was revealed in one APS1 patient, together with autoantibodies against gonadotropes. Autonomic neuropathy was detected in 5 of 10 patients, associated with autoantibodies to adrenal medulla in 2 cases. Of 5 patients with autoantibodies to cerebellar neurons, 2 reported emotional or memory alterations. Conclusions. The majority of Sardinian APS1 patients developed autoantibodies to an assortment of neuroendocrine cells. Novel targets of clinical relevance may include pituitary hormones, uncharacterized pituitary targets, and adrenal medullary cells. An high prevalence of GH deficiency, and possibly of autonomic neuropathy, were also revealed. [ABSTRACT FROM AUTHOR]

Ample animal studies demonstrate that neuropeptides NPY and α-MSH expressed in Arcuate Nucleus and Nucleus of the Tractus Solitarius, modulate glucose homeostasis and food intake. In contrast is the absence of data validating these observations for human disease. Here we compare the post mortem immunoreactivity of the metabolic neuropeptides NPY, αMSH and VGF in the infundibular nucleus, and brainstem of 11 type-2 diabetic and 11 non-diabetic individuals. α-MSH, NPY and tyrosine hydroxylase in human brain are localized in the same areas as in rodent brain. The similar distribution of NPY, α-MSH and VGF indicated that these neurons in the human brain may share similar functionality as in the rodent brain. The number of NPY and VGF immuno positive cells was increased in the infundibular nucleus of diabetic subjects in comparison to non-diabetic controls. In contrast, NPY and VGF were down regulated in the Nucleus of the Tractus Solitarius of diabetic patients. These results suggest an activation of NPY producing neurons in the arcuate nucleus, which, according to animal experimental studies, is related to a catabolic state and might be the basis for increased hepatic glucose production in type-2 diabetes. [ABSTRACT FROM AUTHOR]

Abstract: The vgf gene (non-acronymic) is induced in vivo by neurotrophins including Nerve Growth Factor (NGF), Brain Derived Growth Factor (BDNF) and Glial Derived Growth Factor (GDNF), by synaptic activity and by homeostatic and other stimuli. Post-translational processing of a single VGF precursor gives raise to a varied multiplicity of neuro-endocrine peptides, some of which are secreted upon stimulation both in vitro and in vivo. Several VGF peptides, accounting for ∼20% of the VGF precursor sequence, have shown biological roles including regulation of food intake, energy balance, reproductive and homeostatic mechanisms, synaptic strengthening, long-term potentiation (LTP) and anti-depressant activity. From a further ∼50% of VGF derive multiple “fragments”, largely identified in the human cerebro-spinal fluid by proteomic studies searching for disease biomarkers. These represent an important starting point for discovery of further VGF products relevant to neuronal brain functions, as well as to neurodegenerative and psychiatric disease conditions. A distinct feature of VGF peptides is their cell type specific diversity in all neuroendocrine organs studied so far. Selective differential profiles are found across the cell populations of pituitary, adrenal medulla and pancreatic islets, and in gastric neuroendocrine as well as some further mucosal cells, and are yet to be investigated in neuronal systems. At the same time, specific VGF peptide/s undergo selective modulation in response to organ or cell population relevant stimuli. Such pattern argues for a multiplicity of roles for VGF peptides, including endocrine functions, local intercellular communication, as well as the possible mediation of intracellular mechanisms. [Copyright &y& Elsevier]

Although vgf gene knockout mice are hypermetabolic, administration of the VGF peptide TLQP-21 itself increased energy consumption. Agonist-antagonist roles are thus suggested for different VGF peptides, and the definition of their tissue heterogeneity is mandatory. We studied the rat stomach using antisera to C- or N-terminal sequences of known or predicted VGF peptides in immunohistochemistry and ELISA. TLQP (rat VGF556-565) peptide/s were most abundant (162±11 pmol/g, mean±S.E.M.) and were brightly immunostained in enterochromaffin-like (ECL) cells and somatostatin cells. A peptide co-eluting with TLQP-21 was revealed in HPLC of gastric and hypothalamic extracts, while the extended TLQP-62 form was restricted to the hypothalamus. Novel PGH (rat VGF422-430) peptide/s were revealed in ghrelin cells, mostly corresponding to low MW forms (0.8-1.5 kDa), while VGF C-terminus peptides were confined to neurons. VGF mRNA was present in the above gastric endocrine cell types, and was prominent in chief cells, in parallel with low-intensity staining for further cleaved products from the C-terminal region of VGF (HVLL peptides: VGF605-614). In swine stomach, a comparable profile of VGF peptides was revealed by immunohistochemistry. When fed and fasted rats were studied, a clear-cut, selective decrease on fasting was observed for TLQP peptides only (162±11 vs 74±5.3 pmol/g, fed versus fasted rats, mean±S.E.M., P≤0.00001). In conclusion, specific VGF peptides appear to be widely represented in different gastric endocrine and other mucosal cell populations. The selective modulation of TLQP peptides suggests their involvement in peripheral neuro-endocrine mechanisms related to feeding responses and/or ECL cell regulation. [ABSTRACT FROM AUTHOR]

• TLQP-21 is hypoglycemic. • Upon hyperglycaemia pancreatic TLQP-21 decreases while plasmatic TLQP-21 increases. • TLQP-21 acts as hormone. The TLQP-21 peptide potentiates glucose-stimulated insulin secretion, hence we investigated its endogenous response to glucose. Fasted mice received intraperitoneal glucose (3 g/kg), or saline (controls), and were sacrificed 30 and 120 min later (4 groups, n = 6/group). We investigated TLQP-21 in pancreas and plasma using immunohistochemistry, enzyme-linked immunosorbent assay (ELISA) and high performance liquid chromatography (HPLC), as well as TLQP-21 receptors (gC1q-R and C3a-R1) expression in pancreas by immunohistochemistry. In pancreas, TLQP-immunoreactivity (TLQP-ir.) was shown in insulin-, glucagon- and somatostatin-containing cells. Upon glucose, TLQP-ir. decreased at 30 min (∼40 % vs. controls), while returning to basal values at 120 min. In all groups, C3a-R1 was localized in ∼50 % of TLQP labelled islet cells (mostly central), while gC1q-R was detected in ∼25 % of TLQP cells (mainly peripheral). HPLC fractions of control pancreas extracts, assessed by ELISA, confirmed the presence of a TLQP-21 compatible-form (∼2.5 kDa MW). In plasma, TLQP-ir. increased at 30 min (∼30 %), with highest concentrations at 120 min (both: p< 0.05 vs. controls), while HPLC fractions showed an increase in the TLQP-21 compatible form. Upon hyperglycaemia, TLQP-21 would be released from islets, to enhance insulin secretion but we cannot exclude an autocrine activity which may regulate insulin storage/secretion. [ABSTRACT FROM AUTHOR]

• The VGF-derived TLQP peptides (TLQPp) regulates energy expenditure. • During fasting, TLQPp were down-regulated in the stomach, pancreas and adrenal gland. • During fasting, TLQPp significantly increased within the blood circulation. The VGF-derived TLQP peptides (TLQPp), a new potential drug target for obesity, are expressed in stomach, pancreas, adrenal gland as well as in adipose tissues, and, when exogenously injected, regulate energy expenditure and food intake. However, it is not clear if these peptides physiologically change in these organs in response to fasting. Rats were subdivided into four groups: (A) fed ad libitum , (B) fed with restrictions (once a day) (C) fast for 48 h and (D) fast for 48 h and then fed 1 h before sacrifice. Immunosorbent assay was used to possibly reveal TLQPp changes upon fasting in plasma as well as in pancreas, adrenal gland, stomach and adipose tissues. In the latter organs, we also measured the levels of the VGF precursor protein while immunohistochemistry was used to investigate the presence of the TLQP-21 receptors. During fasting, TLQPp were down-regulated in the stomach (45 %), pancreas (47 %), adrenal gland (51 %) and WAT (45.2 %) in parallel with a significant increase in the blood (36.6 %), all versus ad libitum group. In the same organs where the TLQPp were decreased upon fasting, the VGF precursor levels were not changed. In ad libitum rats, TLQP-21 receptors were well represented within the same cells that expressed TLQPp, suggesting an autocrine activity to be better investigated. During fasting, TLQPp are probably produced and immediately secreted into the blood circulation, until the hypoglycaemia is counteracted. [ABSTRACT FROM AUTHOR]

In a previous proteomic study, we identified the neurosecretory protein VGF (VGF) as a potential biomarker for dementia with Lewy bodies (DLB). Here, we extended the study of VGF by comparing levels in cerebrospinal fluid (CSF) from 44 DLB patients, 20 Alzheimer's disease (AD) patients, and 22 cognitively normal controls selected from the Amsterdam Dementia Cohort. CSF was analyzed using two orthogonal analytical methods: (1) In-house-developed quantitative ELISA and (2) selected reaction monitoring (SRM). We further addressed associations of VGF with other CSF biomarkers and cognition. VGF levels were lower in CSF from patients with DLB compared to either AD patients or controls. VGF was positively correlated with CSF tau and α-synuclein (0.55 < r < 0.75), but not with Aβ1-42. In DLB patients, low VGF levels were related to a more advanced cognitive decline at time of first presentation, whereas high levels of VGF were associated with steeper subsequent longitudinal cognitive decline. Hence, CSF VGF levels were lower in DLB compared to both AD and controls across different analytical methods. The strong associations with cognitive decline further points out VGF as a possible disease stage or prognostic marker for DLB. [ABSTRACT FROM AUTHOR]

While the VGF-derived TLQP peptides have been shown to prevent neuronal apoptosis, and to act on synaptic strengthening, their involvement in Amyotrophic Lateral Sclerosis (ALS) remains unclarified. We studied human ALS patients’ plasma (taken at early to late disease stages) and primary fibroblast cultures (patients vs controls), in parallel with SOD1-G93A transgenic mice (taken at pre-, early- and late symptomatic stages) and the mouse motor neuron cell line (NSC-34) treated with Sodium Arsenite (SA) to induce oxidative stress. TLQP peptides were measured by enzyme-linked immunosorbent assay, in parallel with gel chromatography characterization, while their localization was studied by immunohistochemistry. In controls, TLQP peptides, including forms compatible with TLQP-21 and 62, were revealed in plasma and spinal cord motor neurons, as well as in fibroblasts and NSC-34 cells. TLQP peptides were reduced in ALS patients’ plasma starting in the early disease stage (14% of controls) and remaining so at the late stage (16% of controls). In mice, a comparable pattern of reduction was shown ( vs wild type), in both plasma and spinal cord already in the pre-symptomatic phase (about 26% and 70%, respectively). Similarly, the levels of TLQP peptides were reduced in ALS fibroblasts (31% of controls) and in the NSC-34 treated with Sodium Arsenite (53% of decrease), however, the exogeneous TLQP-21 improved cell viability (SA-treated cells with TLQP-21, vs SA-treated cells only: about 83% vs. 75%). Hence, TLQP peptides, reduced upon oxidative stress, are suggested as blood biomarkers, while TLQP-21 exerts a neuroprotective activity. [ABSTRACT FROM AUTHOR]

Abstract: The effect of NERP-1 and NERP-2, two recently discovered VGF-derived peptides, on feeding and penile erection was studied after injection into the lateral ventricles, the lateral hypothalamus, the arcuate nucleus or the paraventricular nucleus of the hypothalamus. NERP-2 (1–5nmol), but not NERP-1 (2–4nmol), increased feeding in a dose-dependent manner when injected into the lateral ventricles or bilaterally into the lateral hypothalamus but not into the arcuate or the paraventricular nucleus. The effect of NERP-2 given into the lateral ventricles was found in the first, but not in the second 60min after treatment, and was antagonized by SB-408124, an orexin-1 receptor antagonist given into the lateral ventricles or the arcuate nucleus, but not into the paraventricular nucleus. However, SB-408124 was unable to reduce NERP-2-induced feeding when injected bilaterally into the lateral hypothalamus before NERP-2 given also bilaterally into the lateral hypothalamus. In contrast, NERP-1, but not NERP-2, induced penile erection in a dose-dependent manner when injected into the lateral ventricles or the arcuate nucleus, but not into the paraventricular nucleus or the lateral hypothalamus. The pro-erectile effect of NERP-1 was not prevented by the prior injection of d(CH2)5Tyr (Me)2-Orn8-oxytocin or SB-408124 into the lateral ventricles. The present results suggest that while NERP-2 facilitates feeding by acting in the lateral hypothalamus, possibly by increasing orexin activity in the arcuate nucleus, NERP-1 facilitates penile erection by acting in the arcuate nucleus with a mechanism not related to orexin or oxytocin. [Copyright &y& Elsevier]

Abstract: VGF or VGF nerve growth factor inducible is a protein that has been found to play a role in regulating energy homeostasis and metabolism. From VGF precursor derive two neuroendocrine regulatory peptides NERP-1 and NERP-2 that, intracerebroventricular (icv) injected, modulate the antidiuretic hormone (ADH) release. Thus, we investigated possible modulations of the NERPs and other VGF peptides (namely VGF C-terminus, TLQP and PGH) in the hypothalamic–pituitary-axis, adrenal gland and plasma upon osmotic stimuli. The latter tissues were studied using water deprived (WD), salt loaded (SL), rehydrated after salt cargo and control rats by immunohistochemistry and immunoenzymatic assays. The high-performance liquid chromatography ensured the endogenous presence of the two NERPs in both plasma and hypothalamus. Upon dehydration, NERP-1 levels increased in the median eminence (M.E.) only, while using SL rats, the values of both NERPs increased in the M.E. and even in the hypothalamus. Conversely, in the blood of WD and SL rats, the levels of NERP-1 and NERP-2 decreased while, using pituitary from both rat groups, levels of NERP-2 increased and those of NERP-1 decreased. Reduction in the VGF C-terminus peptide levels was observed exclusively in the M.E. (using WD rats) and pituitary (using WD and SL rats), while PGH and TLQP peptide levels never changed in all tissues tested. By immunohistochemistry, the VGF peptides studied (apart from the TLQP peptides) were present in the hypothalamic and pituitary ADH containing neurons of the control rats, while using WD and SL rats, an immunostaining increase was selectively revealed for VGF C-terminus peptides in the hypothalamic neurons that produce ADH. All VGF changes found using SL rats disappeared after only 1h of rehydration. In conclusion, we hypothesize that NERPs may be involved in both autocrine and endocrine mechanisms important for the fluid balance. [Copyright &y& Elsevier]

Abstract: Oxytocin (100 ng) induces penile erection when injected unilaterally into the ventral subiculum of the hippocampus of male rats. The pro-erectile effect started mostly 30 min after treatment and occurred 15 min after an increase in both nitric oxide (NO) production, measured by the concentration of NO2 − and NO3 −, the main metabolites of newly formed NO, and extra-cellular glutamic acid concentration in the dialysate obtained from the ventral subiculum by intracerebral microdialysis. These responses were abolished by d(CH2)5Tyr(Me)2-Orn8-vasotocin (2 μg), an oxytocin receptor antagonist, S-methyl-l-thiocitrulline (SMTC), a selective inhibitor of neuronal NO-synthase (25 μg), and haemoglobin, a NO scavenger (25 μg), given into the ventral subiculum before oxytocin. Unlike d(CH2)5Tyr(Me)2-Orn8-vasotocin, SMTC and haemoglobin, (+)MK-801 (5 μg), a noncompetitive antagonist of NMDA receptors abolished oxytocin-induced penile erection, but reduced only partially the increase in NO production and extra-cellular glutamic acid. As NMDA (0.25–1 μg) injected into the ventral subiculum induces penile erection episodes, which also occurred with an increase of NO production and extra-cellular glutamic acid, and NMDA responses were abolished by (+)MK-801 (5 μg), but not by SMTC (25 μg) or haemoglobin (25 μg), injected into the ventral subiculum, these results show that oxytocin injected into the ventral subiculum increases NO production by activating its own receptors. NO in turn increases glutamic acid neurotransmission, leading to penile erection, possibly through neural (glutamatergic) efferent projections from the ventral subiculum to extra-hippocampal brain areas (e.g., prefrontal cortex) modulating the activity of mesolimbic dopaminergic neurons. [Copyright &y& Elsevier]

B Results: b In the sample of 313 BD patients and 316 controls, partial correlation test corrected for age confirmed the positive correlation between LTL and Li treatment duration in patients with long-term treatment (n = 173, beta = 0.17, p = 0.03). These findings were further supported by preliminary results from a subsample of the cross-diagnostic cohort showing longer LTL in patients with BD treated with Li compared to both MDD and SCZ patients (p = 0.008 and p<0.0001, respectively). B Discussion: b Our data further support previous findings showing that long-term Li treatment has a protective effect against telomere shortening in BD, while patients with SCZ seem to be more susceptible to accelerated telomere shortening. [Extracted from the article]