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Systemic sclerosis (SSc) is an autoimmune disease that affects skin and multiple internal organs. TGF-β, a central trigger of cutaneous fibrosis, activates fibroblasts with the involvement of the stress-inducible chaperone heat shock protein 90 isoform α (Hsp90α). Available evidence supports overexpression and secretion of Hsp90α as a feature in profibrotic pathological conditions. The aim of this work is to investigate the expression and function of Hsp90α in experimental models of skin fibrosis such as human fibroblasts, C57BL/6 mice, and in human SSc. For this purpose, we generated a new experimental model based on doxorubicin administration with improved characteristics with respect to the bleomycin model. We visualized disease progression in vivo by fluorescence imaging. In this work, we obtained Hsp90α mRNA overexpression in human skin fibroblasts, in bleomycin- and doxorubicin-induced mouse fibrotic skin, and in lungs of bleomycin- and doxorubicin-treated mice. Hsp90α-deficient mice showed significantly decreased skin thickness compared with wild-type mice in both animal models. In SSc patients, serum Hsp90α levels were increased in patients with lung involvement and in patients with the diffuse form of SSc (dSSc) compared with patients with the limited form of SSc. The serum Hsp90α levels of patients dSSc were correlated with the Rodnan score and the forced vital capacity variable. These results provide new supportive evidence of the contribution of the Hsp90α isoform in the development of skin fibrosis. In SSc, these results indicated that higher serum levels were associated with dSSc and lung fibrosis., This work was supported by Spanish Ministerio de Economía, Industria y Competitividad, Gobierno de España Grant RTI2018-095214-B-I00, as well as by the Instituto de Formación e Investigación Marqués de Valdecilla IDIVAL (InnVal 17/22; InnVal 20/34), 2020UCI22-PUB-0003 Gobierno de Cantabria (to A.V.V.), SAF2016-75195-R (to J.M.), SAF2017-82905-R (to R.M.), and (NextVal 18/14) to A.P.

Rapid cryopreservation of biological specimens is the gold standard for visualizing cellular structures in their true structural context. However, current commercial cryo-fluorescence microscopes are limited to low resolutions. To fill this gap, we have developed cryoSIM, a microscope for 3D super-resolution fluorescence cryo-imaging for correlation with cryo-electron microscopy or cryo-soft X-ray tomography. We provide the full instructions for replicating the instrument mostly from off-the-shelf components and accessible, user-friendly, open-source Python control software. Therefore, cryoSIM democratizes the ability to detect molecules using super-resolution fluorescence imaging of cryopreserved specimens for correlation with their cellular ultrastructure. Funding: Wellcome Trust (091911/Z/11/Z, 096144/Z/11/Z, 105605/Z/14/Z, 107457/Z/15/Z, 203141/Z/16/Z, 209412/Z/17/Z); H2020Marie Skłodowska-Curie Actions (700184).

To evaluate the impact of COVID-19 lockdown on glycaemic control and diabetes follow-up in a Spanish metropolitan area with a total general population of 340,000.A retrospective real-world study comparing HbA1c testing, an indicator of diabetes control, and mean HbA1c during different COVID-19 restriction periods in 2020 (full lockdown, post-lockdown, partial lockdown) with the same periods in 2019. HbA1c testing was analysed per study period and according to gender, age and clinical setting. Associations between HbA1c testing and different covariables were investigated using logistic regression analysis. Changes in HbA1c were evaluated by repeated measures multivariate analysis of variance (ANOVA).During full lockdown, 6847 individuals, of which 56.7% were over 65 and 6.5% below 40, were tested for HbA1c compared to 14,180 in 2019 (OR 0.47, 95% CI:0.46-0.49). Reduction in HbA1c testing was greater among older individuals (OR 0.44, 95% CI:0.42-0.45). No differences were observed for post-lockdown (OR 1.01, 95% CI:0.99-1.04). During partial lockdown, 10,816 individuals had at least one HbA1c measured compared to 12,749 in 2019 (OR 0.84, 95% CI:0.82-0.87). Mean HbA1c during full lockdown was 7.26% (±1.06) compared to 7.50% (±1.14) in 2019 (p.0001). For gender and across all age groups, HbA1c levels were lower during full lockdown. HbA1c changes were not significantly different during post-lockdown and partial lockdown.COVID-19 restriction measures affected HbA1c testing. During complete lockdown, HbA1c testing decreased by half across all gender and age groups. No deleterious effect on glycaemic control was observed during lockdown and post-lockdown among those tested.

SummaryWhile post-transcriptional control is thought to be required at the periphery of neurons and glia, its extent is unclear. Here, we investigate systematically the spatial distribution and expression of mRNA at single molecule sensitivity and their corresponding proteins of 200 YFP trap protein trap lines across the intact Drosophila nervous system. 98% of the genes studied showed discordance between the distribution of mRNA and the proteins they encode in at least one region of the nervous system. These data suggest that post-transcriptional regulation is very common, helping to explain the complexity of the nervous system. We also discovered that 68.5% of these genes have transcripts present at the periphery of neurons, with 9.5% at the glial periphery. Peripheral transcripts include many potential new regulators of neurons, glia and their interactions. Our approach is applicable to most genes and tissues and includes powerful novel data annotation and visualisation tools for post-transcriptional regulation.Brief outlineA novel high resolution and sensitive approach to systematically co-visualise the distribution of mRNAs and proteins in the intact nervous system reveals that post-transcriptional regulation of gene expression is very common. The rich data landscape is provided as a browsable resource (link), using Zegami, a cloud-based data exploration platform (link). Our solution provides a paradigm for the characterisation of post-transcriptional regulation of most genes and model systems.Highlights196/200 (98%) Drosophila genes show discordant RNA and protein expression in at least one nervous system region137/200 (68.5%) mRNAs are present in at least one synaptic compartmentNovel localised mRNA and protein discovered in periphery of glial processesNew paradigm for analysis of post-transcriptional regulation and data exploration

Objective: To evaluate real-world efficacy and safety of SGLT2 inhibitor (SGLT2i) use in combination with insulin in people with type 1 diabetes. Research design and methods: We conducted a retrospective cohort European two-center study. Data on demographics, HbA1c, weight, insulin use, renal function, and adverse events were collected from 199 adults with type 1 diabetes who initiated a SGLT2i adjunct to insulin. Subgroup analyses were performed to identify who benefited most and who was more at risk for adverse events. Results: Overall, significant reductions in HbA1c (-0.5%), weight (-2.9 kg), and mean daily insulin (-8.5%) were achieved after 12 months. The highest reduction in HbA1c was obtained in individuals with baseline HbA1c >8% (-0.7%) [64mmol/mol]. The largest weight loss was observed in subjects with BMI >27 kg/m2 (-3.5 kg). Individuals with baseline eGFR 2), whereas those with UACR >15 mg/g showed a decrease in UACR (-16.6 mg/g). Fifty-seven individuals (28.6%) reported adverse events: 45 genital infections (22.6%), 5 ketosis episodes (2.5%), and 7 cases of diabetic ketoacidosis (DKA) (3.5%). No severe hypoglycemia events were reported. Conclusions: Our real-world data on SGLT2i showed promising results in reductions in HbA1c, weight, and insulin requirements in type 1 diabetes. Benefits were more pronounced in individuals with higher baseline HbA1c and BMI. DKA remained a major concern, despite educational measures. Further real-life evidence is still required to evaluate SGLT2i longer-term effects and their impact on reno-cardiovascular outcomes.

OBJECTIVE To evaluate real-world efficacy and safety of sodium–glucose cotransporter 2 inhibitor (SGLT2i) use in combination with insulin in people with type 1 diabetes. RESEARCH DESIGN AND METHODS We conducted a retrospective cohort European two-center study. Data on demographics, HbA1c, weight, insulin use, renal function, and adverse events were collected for 199 adults with type 1 diabetes who initiated a SGLT2i adjunct to insulin. Subgroup analyses were performed to identify who benefited most and who was more at risk for adverse events. RESULTS Overall, significant reductions in mean HbA1c (−0.5%), weight (−2.9 kg), and daily insulin (−8.5%) were achieved after 12 months. The greatest reduction in HbA1c was obtained in individuals with baseline HbA1c >8% (−0.7% [64 mmol/mol]). The most weight loss was observed in subjects with BMI >27 kg/m2 (−3.5 kg). Individuals with baseline estimated glomerular filtration rate (eGFR) 15 mg/g showed a decrease in UACR (−16.6 mg/g). Fifty-seven individuals (28.6%) reported adverse events: 45 with genital infections (22.6%), 5 ketosis episodes (2.5%), and 7 diabetic ketoacidosis (DKA) (3.5%). No severe hypoglycemia events were reported. CONCLUSIONS Our real-world data on SGLT2i showed promising results in reductions in HbA1c, weight, and insulin requirements in type 1 diabetes. Benefits were more pronounced in individuals with higher baseline HbA1c and BMI. DKA remained a major concern, despite educational measures. Further real-life evidence is still required for evaluation of SGLT2i longer-term effects and their impact on reno-cardiovascular outcomes.

Thyroid cancer is the most common endocrine malignancy and exhibits rising incidence. Annual incidence varies by sex, age, and geographical location. It has been reported that impairment of vitamin D signalling promotes thyroid cancer progression. Recent studies have shown that vitamin D, a fat-soluble vitamin that acts as both a nutrient and a hormone, may have utility in the prevention of autoimmune thyroid-related diseases. However, the precise role of vitamin D in the pathobiology of thyroid cancer is controversial. Previous studies have suggested that elevated serum vitamin D levels have a protective role in thyroid cancer. However, there is also evidence demonstrating no inverse relationship between vitamin D levels and the occurrence of thyroid cancer. Furthermore, recent data provide evidence that circulating vitamin D concentration is inversely correlated with disease aggressiveness and poor prognosis, while evidence of an association with tumour initiation remains weak. Nevertheless, a variety of data support an anti-tumorigenic role of vitamin D and its potential utility as a secondary chemopreventive agent. In this review, we highlighted recent findings regarding the association of vitamin D status with the risk of thyroid cancer, prognosis, potential mechanisms, and possible utility as a chemopreventive agent.

© 2021 The Author(s)., Protein-based hybrid nanomaterials have recently emerged as promising platforms to fabricate tailored multifunctional biologics for biotechnological and biomedical applications. This work shows a simple, modular, and versatile strategy to design custom protein hybrid nanomaterials. This approach combines for the first time the engineering of a therapeutic protein module with the engineering of a nanomaterial-stabilizing module within the same molecule, resulting in a multifunctional hybrid nanocomposite unachievable through conventional material synthesis methodologies. As the first proof of concept, a multifunctional system was designed ad hoc for the therapeutic intervention and monitoring of myocardial fibrosis. This hybrid nanomaterial combines a designed Hsp90 inhibitory domain and a metal nanocluster stabilizing module resulting in a biologic drug labelled with a metal nanocluster. The engineered nanomaterial actively reduced myocardial fibrosis and heart hypertrophy in an animal model of cardiac remodeling. In addition to the therapeutic effect, the metal nanocluster allowed for in vitro, ex vivo, and in vivo detection and imaging of the fibrotic disease under study. This study evidences the potential of combining protein engineering and protein-directed nanomaterial engineering approaches to design custom nanomaterials as theranostic tools, opening up unexplored routes to date for the next generation of advanced nanomaterials in medicine., This work was partially supported by the European Research Council ERC-CoG-648071-ProNANO and ERC-PoC-841063-NIMM; Agencia Estatal de Investigación, Spain (PID2019-111649RB-I00; RTI2018-095214-B-I00; BIO2016-77367-C2-1-R; and BIO2015-72124-EXP); the Basque Government (Elkartek KK-2017/00008; RIS3-2019222005); and IDIVAL InnVal 17/22. C. S. C. thanks Gipuzkoa Foru Aldundia (Gipuzkoa Fellows program; grant number 2019-FELL-000018-01/62/2019) for financial support. E.L.-M. thanks the Spanish Ministry of Science and Innovation for the FPI grant (BES-2017-079646). This work was performed under the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency – Grant No. MDM-2017-0720 (CIC biomaGUNE). SXRF maps were acquired at the I18 beamline in the frame of the Diamond proposals SP20603.

The lack of an effective, simple, and highly sensitive protocol for fluorescent in situ hybridization (FISH) at the Drosophila larval neuromuscular junction (NMJ) has hampered the study of mRNA biology. Here, we describe our modified single molecule FISH (smFISH) methods that work well in whole mount Drosophila NMJ preparations to quantify primary transcription and count individual cytoplasmic mRNA molecules in specimens while maintaining ultrastructural preservation. The smFISH method is suitable for high-throughput sample processing and 3D image acquisition using any conventional microscopy imaging modality and is compatible with the use of antibody colabeling and transgenic fluorescent protein tags in axons, glia, synapses, and muscle cells. These attributes make the method particularly amenable to super-resolution imaging. With 3D Structured Illumination Microscopy (3D-SIM), which increases spatial resolution by a factor of 2 in X, Y, and Z, we acquire super-resolution information about the distribution of single molecules of mRNA in relation to covisualized synaptic and cellular structures. Finally, we demonstrate the use of commercial and open source software for the quality control of single transcript expression analysis, 3D-SIM data acquisition and reconstruction as well as image archiving management and presentation. Our methods now allow the detailed mechanistic and functional analysis of sparse as well as abundant mRNAs at the NMJ in their appropriate cellular context.

BACKGROUND Diabetic peripheral neuropathy (DPN) is a chronic complication of diabetes mellitus associated with high morbidity and mortality. Major risk factors for DPN include metabolic changes, duration of diabetes, nerve ischaemia and derangements in regeneration and nerve repair programmes. Chemokines have been previously implicated in the pathogenesis of various neuropathies and neuropathic pain processes. The aim of this pilot study was to evaluate the association between the plasma levels of chemokines (CXCL9, CXCL10 and CXCL11) in the presence of DPN in a cohort of type 2 diabetes (T2D) patients. MATERIALS AND METHODS We studied 73 patients with T2D: 36 with DPN and 37 without DPN. DPN was established through the Semmes-Weinstein test (SW). Plasma levels of circulating chemokines CXCL9, CXCL10 and CXCL11 were determined using DuoSet ELISA kits (Abingdon, UK). RESULTS We found that levels of CXCL10 were significantly higher in patients with DPN than amongst patients without DPN (57.6 ± 38.3 vs 38.1 ± 33.4 pg/mL, respectively; P = .034). Serum levels of chemokine CXCL9 were also higher amongst patients with DPN but did not reach a statistical significance (188.1 ± 72.7 and 150.4 ± 83.6 pg/mL, respectively, P = .06). CONCLUSIONS Increased circulating levels of CXCL10 were associated with DPN in T2D patients, suggesting a role of this chemokine in the DPN. Determination of CXCL10 levels could be used as a marker for the early detection and implementation of therapeutic strategies in order to reverse and prevent the DPN.

Cardiac fibrosis is a pathological process that presents a continuous overproduction of extracellular matrix (ECM) components in the myocardium, which negatively influences the progression of many cardiac diseases. Transforming growth factor β (TGFβ) is the main ligand that triggers the production of pro-fibrotic ECM proteins. In the cardiac fibrotic process, TGFβ and its canonical signaling mediators are tightly regulated at different levels as well as epigenetically. Cardiac fibroblasts are one of the most important TGFβ target cells activated after cardiac injury. TGFβ-driven fibroblast activation is subject to epigenetic modulation and contributes to the progression of cardiac fibrosis, mainly through the expression of pro-fibrotic molecules implicated in the disease. In this review, we describe epigenetic regulation related to canonical TGFβ signaling in cardiac fibroblasts., This work was supported by the Agencia Estatal de Investigación, Spain (RTI2018-095214-B-I00; and BIO2015-72124-EXP), and the IDIVAL InnVal 17/22, NextVal 18/11 Thanks to the University of Cantabria and the IDIVAL (UC-IDIVAL Fellows program; grant number PREVAL19/05: Maestro D and UC: Algeciras L) for financial support.

Background : Diabetic peripheral neuropathy (DPN) has high morbidity and mortality. Major risk factors of DPN include metabolic changes, duration of diabetes, nerve ischemia, and derangements in regeneration and nerve repair programs. The implication of chemokines in the pathogenesis of various neuropathies and neuropathic pain processes has been studied previously. This pilot study aimed to evaluate the association between plasma levels of chemokines CXCL9, CXCL10, and CXCL11 with the presence of DPN in a cohort of type 2 diabetes (T2D) patients. Methods : We have studied a total of 73 T2D patients (36 patients with DPN, and 37 without DPN). DPN was established through the Semmes-Weinstein test (SW). Plasma levels of circulating chemokines CXCL9, CXCL10, and CXCL11 were determined using Duoset ELISA kits (Abingdon, UK). Results: In T2D patients with similar gender distribution, duration of the disease, peripheral macroangiopathy, and metabolic control, we detected significantly higher serum levels of chemokine CXCL10 among patients with DPN than among patients without DPN (57.6 ± 38.3 vs. 38.1 ± 33.4 pg/mL, respectively; p = 0.034). Serum levels of chemokine CXCL9 were also higher among patients with DPN (188.1 ± 72.7 and 150.4 ± 83.6 pg/mL, respectively, p = 0.06). Conclusions : DPN in T2D patients was associated with a significant increase in CXCL10 circulating levels, suggesting a role for this chemokine in the DPN. Novel inflammatory markers that allow for early detection and therapeutic strategies in order to reverse and prevent the DPN should be investigated.

Rapid cryo-preservation of biological specimens is the gold standard for visualising cellular structures in their true structural context. However, current commercial cryo-fluorescence microscopes are limited to low resolutions. To fill this gap, we have developed cryoSIM, a microscope for 3D super-resolution fluorescence cryo-imaging for correlation with cryo electron microscopy or cryo soft X-ray tomography. We provide the full instructions for replicating the instrument mostly from off-the-shelf components and accessible, user-friendly open source Python control software. Therefore, cryoSIM democratises the ability to detect molecules using super-resolution fluorescence imaging of cryo-preserved specimens for correlation with their cellular ultrastructure.

Aims To analyze therapeutic inertia in type 2 diabetes (T2D) subjects with suboptimal glycemic control and treated with ≥2 non-insulin antidiabetic agents in a primary care setting. Methods A retrospective study was conducted using electronic medical records from subjects with HbA1c ≥7.0% (≥53 mmol/mol). Therapeutic inertia was defined as the absence of treatment intensification despite suboptimal glycemic control where intensification should have been implemented (HbA1c ≥7.5% [≥58 mmol/mol]). Time to the first intensification with non-insulin antidiabetic agent or insulin and HbA1c values at the time of intensification were evaluated by competing risk analysis. Results 2652 adults with T2D and HbA1c ≥7.0% (≥53 mmol/mol) were included. During the 4-year follow-up, among 1628 individuals with HbA1c ≥7.5% [≥58 mmol/mol], therapeutic inertia was present in 42.9% of cases. Median time to intensification was 14.5 months (IQR25–75, 4–24 months). In this subgroup, 72.7% of subjects initiated non-insulin agents whereas 27.3% initiated insulin. Mean HbA1c values at initiation of treatment intensification were 8.6% (70 mmol/mol) and 9.2% (77 mmol/mol), respectively. Conclusions Therapeutic inertia occurred in over 40% of subjects. Treatment intensification took longer and was performed at higher HbA1c than recommended in clinical guidelines. Reducing therapeutic inertia is a priority to achieve therapeutic goals and prevent chronic complications in T2D.

Neurons are highly vulnerable to genotoxic agents. To restore genome integrity upon DNA lesions, neurons trigger a DNA damage response (DDR) that requires chromatin modifications and transcriptional silencing at DNA damage sites. To study the reorganization of the active RNA polymerase II (Pol II), which transcribes all mRNA-encoding genes, and the participation of the ubiquitin-proteasome system (UPS) in the neuronal DDR, we have used rat sensory ganglion neurons exposed to X-rays (4 Gy) ionizing radiation (IR). In control neurons, Pol II appears concentrated in numerous chromatin microfoci identified as transcription factories by the incorporation of 5'-fluorouridine into nascent RNA. Upon IR treatment, numerous IR-induced foci (IRIF), which were immunoreactive for γH2AX and 53BP1, were observed as early as 30 min post-IR; their number progressively reduced at 3 h, 1 day, and 3 days post-IR. The formation of IRIF was associated with a decrease in Pol II levels by both immunofluorescence and Western blotting. Treatment with the proteasome inhibitor bortezomib strongly increased Pol II levels in both control and irradiated neurons, suggesting that proteasome plays a proteolytic role by clearing stalled Pol II complexes at DNA damage sites, as a prelude to DNA repair. Neuronal IRIF recruited ubiquitylated proteins, including ubiquitylated histone H2A (Ub-H2A), and the catalytic proteasome 20S. Ub-H2A has been associated with transcriptional silencing at DNA damage sites. On the other hand, the participation of UPS in neuronal DDR may be essential for the ubiquitylation of Pol II and other proteasome substrates of the DNA repair machinery and their subsequent proteasome-mediated degradation.

Background and aims: Cardiovascular disease is the leading cause of morbidity and mortality in patients with chronic kidney disease (CKD) and diabetes. Traditional cardiovascular risk factors fail to fully account for the increase in cardiovascular risk in these patients. This study aims to analyse the prevalence and progression of subclinical atherosclerosis in CKD patients with and without diabetes. Methods: We included data from CKD patients with and without diabetes free from previous cardiovascular events from the NEFRONA cohort. Patients underwent baseline and 24-month follow-up carotid and femoral ultrasound examinations. Multivariable models were used to assess the contribution of diabetes to the presence and plaque progression. Results: A total of 419 patients with diabetes and 1129 without diabetes were included. Diabetic patients were older, had higher BMIs, more hypertension and dyslipidaemia. At baseline, the proportion of patients with plaque was higher among diabetic patients (81.4% vs. 64.1%, p < 0.001). Diabetic patients more frequently had more than two vascular territories with plaque (64.4% vs. 48.4%, p < 0.001). Multivariable analysis indicated that plaque at baseline was significantly associated with age, gender, smoking and renal replacement therapy (RRT) in the non-diabetic patients, but only with age and male gender in diabetic patients. Plaque progression was significantly associated with age, number of territories with basal plaque, smoking and RRT in both groups. Conclusions: Subclinical atherosclerosis is more prevalent, carries a higher plaque burden and is more rapidly progressive in renal patients with diabetes. In these patients, diabetes outweighs other described risk factors associated with the presence of subclinical atherosclerosis. (c) 2018 Elsevier B.V. All rights reserved.

Chronic alcohol consumption may cause neurodevelopmental and neurodegenerative disorders. Alcohol neurotoxicity is associated with the production of acetaldehyde and reactive oxygen species that induce oxidative DNA damage. However, the molecular mechanisms by which ethanol disturbs the DNA damage response (DDR), resulting in a defective DNA repair, remain unknown. Here, we have used cultured primary cortical neurons exposed to 50 or 100 mM ethanol for 7 days to analyze the ethanol-induced DDR. Ethanol exposure produced a dose-dependent generation of double strand breaks and the formation of DNA damage foci immunoreactive for the histone gamma H2AX, a DNA damage marker, and for the ubiquitylated H2A, which is involved in chromatin remodeling at DNA damage sites. Importantly, these DNA damage foci failed to recruit the protein 53BP1, a crucial DNA repair factor. This effect was associated with a drop in 53BP1 mRNA and protein levels and with an inhibition of global transcription. Moreover, ethanol-exposed neurons treated with ionizing radiation (2 Gy) also failed to recruit 53BP1 at DNA damage foci and exhibited a greater vulnerability to DNA lesions than irradiated control neurons. Our results support that defective DNA repair, mediated by the deficient expression and recruitment of 53BP1 to DNA damage sites, represents a novel mechanism involved in ethanol neurotoxicity. The design of therapeutic strategies that increase or stabilize 53BP1 levels might potentially promote DNA repair and partially compensate alcohol neurotoxicity.

Cajal bodies (CBs) are nuclear organelles involved in the maturation of spliceosomal small nuclear ribonucleoproteins (snRNPs). They concentrate coilin, snRNPs and the survival motor neuron protein (SMN). Dysfunction of CB assembly occurs in spinal muscular atrophy (SMA). Here, we demonstrate that SMN is a SUMO1 target that has a small ubiquitin-related modifier (SUMO)-interacting motif (SIM)-like motif in the Tudor domain. The expression of SIM-like mutant constructs abolishes the interaction of SMN with the spliceosomal SmD1 (also known as SNRPD1), severely decreases SMN-coilin interaction and prevents CB assembly. Accordingly, the SMN SIM-like-mediated interactions are important for CB biogenesis and their dysfunction can be involved in SMA pathophysiology. © 2014. Published by The Company of Biologists Ltd., Dirección General de Investigación of Spain [grant number BFU2011-23983].

The conversion of muscle into meat is a complex process of major concern for meat scientists due to its influence on the final meat quality. The aim of this study was to investigate the occurrence of autophagic processes in the conversion of muscle into meat. Our findings demonstrated, for the first time, the occurrence of autophagic processes in the muscle tissue at early postmortem period (2 h to 24 h) in both beef breeds studied (Asturiana de los Valles and Asturiana de la Montana) showing significant time-scale differences between breeds, which could indicate a role of this process in meat maturation. These breeds have different physiological features: while Asturiana de los Valles is a meat-specialized breed showing high growth rate, an elevated proportion of white fibers in the muscle and low intramuscular fat level, Asturiana de la Montana is a small- to medium-sized rustic breed adapted to less-favored areas, showing more red fibers in the muscle and a high intramuscular fat content.

Bortezomib is a reversible proteasome inhibitor used as an anticancer drug. However, its clinical use is limited since it causes peripheral neurotoxicity. We have used Sprague–Dawley rats as an animal model to investigate the cellular mechanisms affected by both short-term and chronic bortezomib treatments in sensory ganglia neurons. Proteasome inhibition induces dose-dependent alterations in the architecture, positioning, shape and polarity of the neuronal nucleus. It also produces DNA damage without affecting neuronal survival, and severe disruption of the protein synthesis machinery at the central cytoplasm accompanied by decreased expression of the brain-derived neurotrophic factor. As a compensatory or adaptive survival response against proteotoxic stress caused by bortezomib treatment, sensory neurons preserve basal levels of transcriptional activity, up-regulate the expression of proteasome subunit genes, and generate a new cytoplasmic perinuclear domain for protein synthesis. We propose that proteasome activity is crucial for controlling nuclear architecture, DNA repair and the organization of the protein synthesis machinery in sensory neurons. These neurons are primary targets of bortezomib neurotoxicity, for which reason their dysfunction may contribute to the pathogenesis of the bortezomib-induced peripheral neuropathy in treated patients.

Type I spinal muscular atrophy (SMA) is an autosomal recessive disorder caused by loss or mutations of the survival motor neuron 1 (SMN1) gene. The reduction in SMN protein levels in SMA leads to degeneration and death of motor neurons. In this study, we have analyzed the nuclear reorganization of Cajal bodies, PML bodies and nucleoli in type I SMA motor neurons with homozygous deletion of exons 7 and 8 of the SMN1 gene. Western blot analysis is is revealed a marked reduction of SMN levels compared to the control sample. Using a neuronal dissociation procedure to perform a careful immunocytochemical and quantitative analysis of nuclear bodies, we demonstrated a severe decrease in the mean number of Cajal bodies per neuron and in the proportion of motor neurons containing these structures in type I SMA. Moreover, most Cajal bodies fail to recruit SMN and spliceosomal snRNPs, but contain the proteasome activator PA28, a molecular marker associated with the cellular stress response. Neuronal stress in SMA motor neurons also increases PML body number. The existence of chromatolysis and eccentric nuclei in SMA motor neurons correlates with Cajal body disruption and nucleolar relocalization of coil in, a Cajal body marker. Our results indicate that the Cajal body is a pathophysiological target in type I SMA motor neurons. They also suggest the Cajal body-dependent dysfunction of snRNP biogenesis and, therefore, pre-mRNA splicing in these neurons seems to be an essential component for SMA pathogenesis.

Neurons are very sensitive to DNA damage induced by endogenous and exogenous genotoxic agents, as defective DNA repair can lead to neurodevelopmental disorders, brain tumors and neurodegenerative diseases with severe clinical manifestations. Understanding the impact of DNA damage/repair mechanisms on the nuclear organization, particularly on the regulation of transcription and cell cycle, is essential to know the pathophysiology of defective DNA repair syndromes. In this work, we study the nuclear architecture and spatiotemporal organization of chromatin compartments involved in the DNA damage response (DDR) in rat sensory ganglion neurons exposed to X-ray irradiation (IR). We demonstrate that the neuronal DDR involves the formation of two categories of DNA-damage processing chromatin compartments: transient, disappearing within the 1 day post-IR, and persistent, where unrepaired DNA is accumulated. Both compartments concentrate components of the DDR pathway, including γH2AX, pATM and 53BP1. Furthermore, DNA damage does not induce neuronal apoptosis but triggers the G0–G1 cell cycle phase transition, which is mediated by the activation of the ATM-p53 pathway and increased protein levels of p21 and cyclin D1. Moreover, the run on transcription assay reveals a severe inhibition of transcription at 0.5 h post-IR, followed by its rapid recovery over the 1 day post-IR in parallel with the progression of DNA repair. Therefore, the response of healthy neurons to DNA damage involves a transcription- and cell cycle-dependent but apoptosis-independent process. Furthermore, we propose that the segregation of unrepaired DNA in a few persistent chromatin compartments preserves genomic stability of undamaged DNA and the global transcription rate in neurons.

The consequences of infection at the site of surgery can be devastating to the patient and costly to manage, often requiring readmission to hospital. Surgical site infections account for 14% of all healthcare acquired infections.1 This article summarises the most recent guidance from the National Institute for Health and Clinical Excellence (NICE) for the prevention and management of surgical site infection.2 NICE recommendations are based on systematic reviews of best available evidence. When minimal evidence is available, recommendations are based on the guideline development group’s opinion of what constitutes good practice. Evidence levels for the recommendations are given in italic in square brackets. ### Information for patients and carers Provide patients and carers with clear and consistent, easily understood information and advice throughout all stages of their care. Information should include: [ All these recommendations are based on a high quality randomised controlled trial and on the opinion of the Guideline Development Group ] ### Preoperative phase #### Showering #### Hair removal

The dysfunction of the ubiquitin proteasome system has been related to a broad array of neurodegenerative disorders in which the accumulation of misfolded protein aggregates causes proteotoxicity. The ability of proteasome inhibitors to induce cell cycle arrest and apoptosis has emerged as a powerful strategy for cancer therapy. Bortezomib is a proteasome inhibitor used as an antineoplastic drug, although its neurotoxicity frequently causes a severe sensory peripheral neuropathy. In this study we used a rat model of bortezomib treatment to study the nucleolar and Cajal body responses to the proteasome inhibition in sensory ganglion neurons that are major targets of bortezomib-induced neurotoxicity. Treatment with bortezomib induced dose-dependent dissociation of protein synthesis machinery (chromatolysis) and nuclear retention of poly(A) RNA granules resulting in neuronal dysfunction. However, as a compensatory response to the proteotoxic stress, both nucleoli and Cajal bodies exhibited reactive changes. These include an increase in the number and size of nucleoli, strong nucleolar incorporation of the RNA precursor 5′-fluorouridine, and increased expression of both 45S rRNA and genes encoding nucleolar proteins UBF, fibrillarin and B23. Taken together, these findings appear to reflect the activation of the nucleolar transcription in response to proteotoxic stress Furthermore, the number of Cajal bodies, a parameter related to transcriptional activity, increases upon proteasome inhibition. We propose that nucleoli and Cajal bodies are important targets in the signaling pathways that are activated by the proteotoxic stress response to proteasome inhibition. The coordinating activity of these two organelles in the production of snRNA, snoRNA and rRNA may contribute to neuronal survival after proteasome inhibition. This article is part of a Special Issue entitled: Role of the Nucleolus in Human Disease.

© 2021 The Author(s)., Revealing the intracellular location of novel therapeutic agents is paramount for the understanding of their effect at the cell ultrastructure level. Here, we apply a novel correlative cryo 3D imaging approach to determine the intracellular fate of a designed protein–nanomaterial hybrid with antifibrotic properties that shows great promise in mitigating myocardial fibrosis. Cryo 3D structured illumination microscopy (cryo-3D-SIM) pinpoints the location and cryo soft X-ray tomography (cryo-SXT) reveals the ultrastructural environment and subcellular localization of this nanomaterial with spatial correlation accuracy down to 70 nm in whole cells. This novel high resolution 3D cryo correlative approach unambiguously locates the nanomaterial after overnight treatment within multivesicular bodies which have been associated with endosomal trafficking events by confocal microscopy. Moreover, this approach allows assessing the cellular response towards the treatment by evaluating the morphological changes induced. This is especially relevant for the future usage of nanoformulations in clinical practices. This correlative super-resolution and X-ray imaging strategy joins high specificity, by the use of fluorescence, with high spatial resolution at 30 nm (half pitch) provided by cryo-SXT in whole cells, without the need of staining or fixation, and can be of particular benefit to locate specific molecules in the native cellular environment in bio-nanomedicine., This project has received funding from the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 754397. This work was partially supported by the European Research Council ERC-CoG-648071-ProNANO, ERC-PoC-841063-NIMM; Spanish State Research Agency, Spain; (BIO2015-72124-EXP), (MDM-2017-0720), (PID2019-111649RB-I00) and (RTI2018-095214-B-I00); the Basque Government (Elkartek KK-2017/00008; RIS3-2019222005); and the Next Generation and Innovation Valdecilla (PREVAL19/05), (IDIVAL INNVAL17/22), (INNVAL20/34), (NVAL18/11) and (NextVal 18/14). This work was performed under the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency – Grant No. MDM-2017-0720 (CIC biomaGUNE).