From cohorts to molecules: Adverse impacts of endocrine disrupting mixtures.

Convergent evidence associates exposure to endocrine disrupting chemicals (EDCs) with major human diseases, even at regulation-compliant concentrations. This might be because humans are exposed to EDC mixtures, whereas chemical regulation is based on a risk assessment of individual compounds. Here, we developed a mixture-centered risk assessment strategy that integrates epidemiological and experimental evidence. We identified that exposure to an EDC mixture in early pregnancy is associated with language delay in offspring. At human-relevant concentrations, this mixture disrupted hormone-regulated and disease-relevant regulatory networks in human brain organoids and in the model organisms Xenopus leavis and Danio rerio, as well as behavioral responses. Reinterrogating epidemiological data, we found that up to 54% of the children had prenatal exposures above experimentally derived levels of concern, reaching, for the upper decile compared with the lowest decile of exposure, a 3.3 times higher risk of language delay.

mTORC1 promotes malignant large cell/anaplastic histology and is a targetable vulnerability in SHH-TP53 mutant medulloblastoma.

Medulloblastoma (MB), one of the most malignant brain tumors of childhood, comprises distinct molecular subgroups, with p53 mutant sonic hedgehog-activated (SHH-activated) MB patients having a very severe outcome that is associated with unfavorable histological large cell/anaplastic (LC/A) features. To identify the molecular underpinnings of this phenotype, we analyzed a large cohort of MB developing in p53-deficient Ptch+/- SHH mice that, unexpectedly, showed LC/A traits that correlated with mTORC1 hyperactivation. Mechanistically, mTORC1 hyperactivation was mediated by a decrease in the p53-dependent expression of mTORC1 negative regulator Tsc2. Ectopic mTORC1 activation in mouse MB cancer stem cells (CSCs) promoted the in vivo acquisition of LC/A features and increased malignancy; accordingly, mTORC1 inhibition in p53-mutant Ptch+/- SHH MB and CSC-derived MB resulted in reduced tumor burden and aggressiveness. Most remarkably, mTORC1 hyperactivation was detected only in p53-mutant SHH MB patient samples, and treatment with rapamycin of a human preclinical model phenocopying this subgroup decreased tumor growth and malignancy. Thus, mTORC1 may act as a specific druggable target for this subset of SHH MB, resulting in the implementation of a stringent risk stratification and in the potentially rapid translation of this precision medicine approach into the clinical setting.

Long-term Inhibition of Soleus H-reflex with Epidural Adhesiolysis and Pulsed Radiofrequency in Lumbosacral Neuropathic Pain.

INTRODUCTION: Scientific data about neurophysiological changes subsequent to pulsed radiofrequency (PRF) are still lacking. The goal of this study was to evaluate sural nerve conduction and Hoffmann reflex (H-reflex) in soleus muscle following adhesiolysis and PRF in patients with unilateral chronic lumbosacral L5-S1 neuropathic radiating pain. METHODS: Seventeen patients received two cycles of 240 seconds high-voltage PRF and epidural adhesiolysis. Sural nerve action potential (SNAP) and the ratio of maximum H-reflex to maximum M response (H/M ratio) as well as pain scores were collected in both lower limbs before, immediately following, and 1 month after the treatment. RESULTS: At follow-up, a significant reduction in numeric rating scale (NRS) and Douleur Neuropathique 4 Questions (DN4) scores was observed in 53% of patients reporting pain improvement of ≥ 30% over baseline. The H/M ratio was decreased in the affected limb following PRF (P = 0.01) and 1 month after the treatment (P = 0.04). A direct correlation was observed between H/M ratio variation and NRS score at follow-up in the treated limb (P = 0.04). No significant difference in sural nerve latency, amplitude, and velocity was detected between affected and normal side after treatment and at follow-up. CONCLUSIONS: Epidural adhesiolysis and PRF of the dorsal root ganglion seem to significantly affect spinal reflexes in patients with lumbosacral neuropathic radiating pain.

High-throughput screening identifies histone deacetylase inhibitors that modulate GTF2I expression in 7q11.23 microduplication autism spectrum disorder patient-derived cortical neurons.

BACKGROUND: Autism spectrum disorder (ASD) is a highly prevalent neurodevelopmental condition affecting almost 1% of children, and represents a major unmet medical need with no effective drug treatment available. Duplication at 7q11.23 (7Dup), encompassing 26-28 genes, is one of the best characterized ASD-causing copy number variations and offers unique translational opportunities, because the hemideletion of the same interval causes Williams-Beuren syndrome (WBS), a condition defined by hypersociability and language strengths, thereby providing a unique reference to validate treatments for the ASD symptoms. In the above-indicated interval at 7q11.23, defined as WBS critical region, several genes, such as GTF2I, BAZ1B, CLIP2 and EIF4H, emerged as critical for their role in the pathogenesis of WBS and 7Dup both from mouse models and human studies. METHODS: We performed a high-throughput screening of 1478 compounds, including central nervous system agents, epigenetic modulators and experimental substances, on patient-derived cortical glutamatergic neurons differentiated from our cohort of induced pluripotent stem cell lines (iPSCs), monitoring the transcriptional modulation of WBS interval genes, with a special focus on GTF2I, in light of its overriding pathogenic role. The hits identified were validated by measuring gene expression by qRT-PCR and the results were confirmed by western blotting. RESULTS: We identified and selected three histone deacetylase inhibitors (HDACi) that decreased the abnormal expression level of GTF2I in 7Dup cortical glutamatergic neurons differentiated from four genetically different iPSC lines. We confirmed this effect also at the protein level. LIMITATIONS: In this study, we did not address the molecular mechanisms whereby HDAC inhibitors act on GTF2I. The lead compounds identified will now need to be advanced to further testing in additional models, including patient-derived brain organoids and mouse models recapitulating the gene imbalances of the 7q11.23 microduplication, in order to validate their efficacy in rescuing phenotypes across multiple functional layers within a translational pipeline towards clinical use. CONCLUSIONS: These results represent a unique opportunity for the development of a specific class of compounds for treating 7Dup and other forms of intellectual disability and autism.

Electrocatheter-mediated High-voltage Pulsed Radiofrequency of the Dorsal Root Ganglion in the Treatment of Chronic Lumbosacral Neuropathic Pain: A Randomized Controlled Study.

OBJECTIVES: Despite the interest in scientific community, there is still poor evidence about pulsed radiofrequency (PRF) efficacy in the treatment of neuropathic pain. In order to determine whether high-voltage PRF and epidural adhesiolysis (PRF-EA) showed better results than epidural adhesiolysis alone (EA), a randomized, double-blind, comparative-effectiveness study was conducted in patients with chronic lumbosacral radiating pain and neuropathic features. MATERIALS AND METHODS: A total of 41 patients were randomly allocated to 2 groups. Twenty-one patients were randomized to receive 2 cycles of 240 seconds high-voltage PRF followed by the injection of local anesthetics, hyaluronidase, and betamethasone, whereas 20 patients underwent sham stimulation followed by adhesiolysis. The treatment was delivered at the affected lumbosacral roots and patients, treating physicians and assessors were blinded to intervention. RESULTS: A significant reduction of radiating pain was observed in mean Numeric Rating Scale score at follow-up. A change of -3.43 versus -1.75 (P=0.031) after 1 month and -3.34 versus -0.80 (P=0.005) after 6 months was reported in patients undergoing PRF-EA in comparison with EA, respectively. After 1 month, 57% of patients in the PRF-EA group experienced a pain reduction of ≥50% versus only 25% of patients allocated to EA (P=0.037). Improvement decreased to 48% in the PRF-EA group whereas only 10% of EA reported significant pain relief after 6 months (P=0.008). DISCUSSION: High-voltage PRF of dorsal root ganglion delivered through multifunctional electrode provided significant pain relief and may be considered a valuable treatment in chronic lumbosacral radicular pain with neuropathic features.

Dosage analysis of the 7q11.23 Williams region identifies BAZ1B as a major human gene patterning the modern human face and underlying self-domestication.

We undertook a functional dissection of chromatin remodeler BAZ1B in neural crest (NC) stem cells (NCSCs) from a uniquely informative cohort of typical and atypical patients harboring 7q11.23 copy number variants. Our results reveal a key contribution of BAZ1B to NCSC in vitro induction and migration, coupled with a crucial involvement in NC-specific transcriptional circuits and distal regulation. By intersecting our experimental data with new paleogenetic analyses comparing modern and archaic humans, we found a modern-specific enrichment for regulatory changes both in BAZ1B and its experimentally defined downstream targets, thereby providing the first empirical validation of the human self-domestication hypothesis and positioning BAZ1B as a master regulator of the modern human face. In so doing, we provide experimental evidence that the craniofacial and cognitive/behavioral phenotypes caused by alterations of the Williams-Beuren syndrome critical region can serve as a powerful entry point into the evolution of the modern human face and prosociality.

Interventional treatment for neuropathic pain due to combined cervical radiculopathy and carpal tunnel syndrome: a case report.

The coexistence of median and cervical nerve root damage might hide a complex pathophysiology. Here, we describe and discuss the case of a patient suffering from numbness and painful tingling of the hand, whose symptoms were effectively treated with pulsed radiofrequency and epidural administration of bupivacaine and morphine.

Combined epidural morphine and bupivacaine in the treatment of lumbosacral radicular neuropathic pain: a noncontrolled prospective study.

OBJECTIVE: The aim of this study was to investigate the therapeutic effectiveness of epidural morphine and bupivacaine in patients with chronic lumbosacral radicular neuropathic pain after the cessation of treatment. METHODS: Twenty-two patients with chronic lumbosacral pain with neuropathic features were enrolled. An indwelling catheter was placed into the epidural space, and each patient received an epidural injection of morphine chlorhydrate and bupivacaine up to three times a day. The medication was administered for 4 weeks. The pain intensity score on a 0-10 numeric rating scale (NRS), the total pain rating index rank (PRIr-T), and its coefficients were evaluated before treatment and 1 month after catheter removal. P-value <0.05 was considered statistically significant. RESULTS: NRS and PRIr-T were significantly reduced at follow-up (P=0.001 and P=0.03, respectively), whereas the parallel evolution of the two scores (r=0.75 and P<0.001, respectively) confirmed significant pain relief lasting up to 1 month after treatment cessation. None of the four pain rating coefficients was significantly modified compared to the others in either responders or nonresponders. Successful clinical outcome (pain reduction >30% in NRS) was reached and maintained in half of the patients at follow-up. CONCLUSION: Combined epidural morphine and bupivacaine seems to be effective in the treatment of neuropathic pain.

TRIC: an automated alignment strategy for reproducible protein quantification in targeted proteomics.

Next-generation mass spectrometric (MS) techniques such as SWATH-MS have substantially increased the throughput and reproducibility of proteomic analysis, but ensuring consistent quantification of thousands of peptide analytes across multiple liquid chromatography-tandem MS (LC-MS/MS) runs remains a challenging and laborious manual process. To produce highly consistent and quantitatively accurate proteomics data matrices in an automated fashion, we developed TRIC (http://proteomics.ethz.ch/tric/), a software tool that utilizes fragment-ion data to perform cross-run alignment, consistent peak-picking and quantification for high-throughput targeted proteomics. TRIC reduced the identification error compared to a state-of-the-art SWATH-MS analysis without alignment by more than threefold at constant recall while correcting for highly nonlinear chromatographic effects. On a pulsed-SILAC experiment performed on human induced pluripotent stem cells, TRIC was able to automatically align and quantify thousands of light and heavy isotopic peak groups. Thus, TRIC fills a gap in the pipeline for automated analysis of massively parallel targeted proteomics data sets.

7q11.23 dosage-dependent dysregulation in human pluripotent stem cells affects transcriptional programs in disease-relevant lineages.

Cell reprogramming promises to make characterization of the impact of human genetic variation on health and disease experimentally tractable by enabling the bridging of genotypes to phenotypes in developmentally relevant human cell lineages. Here we apply this paradigm to two disorders caused by symmetrical copy number variations of 7q11.23, which display a striking combination of shared and symmetrically opposite phenotypes--Williams-Beuren syndrome and 7q-microduplication syndrome. Through analysis of transgene-free patient-derived induced pluripotent stem cells and their differentiated derivatives, we find that 7q11.23 dosage imbalance disrupts transcriptional circuits in disease-relevant pathways beginning in the pluripotent state. These alterations are then selectively amplified upon differentiation of the pluripotent cells into disease-relevant lineages. A considerable proportion of this transcriptional dysregulation is specifically caused by dosage imbalances in GTF2I, which encodes a key transcription factor at 7q11.23 that is associated with the LSD1 repressive chromatin complex and silences its dosage-sensitive targets.

Effectiveness of pulsed radiofrequency with multifunctional epidural electrode in chronic lumbosacral radicular pain with neuropathic features.

BACKGROUND: Lumbosacral radicular pain is a common clinical finding with a statistical prevalence ranging from 9.9% to 25% in the general population. OBJECTIVE: To investigate the effectiveness of dorsal root ganglion pulsed radiofrequency (PRF) in patients with chronic lumbosacral radicular pain and neuropathic features. STUDY DESIGN: Prospective case series clinical outcome study. METHODS: We evaluated 34 patients with lumbosacral neuropathic pain who underwent PRF at the corresponding level of radicular symptoms distribution (ranging from L3 to S1). Each patient suffered a single leg-radiating pain with probable neuropathic features (assessed with clinical grading) lasting for > 6 months and unresponsive to previous treatments. A multifunctional PASHA-electrode® was introduced with trans-sacral access through a hollow needle, placed under fluoroscopic guidance into the lumbosacral epidural space and its active tip moved close to the dorsal root ganglion responsible of the clinical symptoms. After connecting the electrode to a generator, stimulation tests were performed and PRF was started and applied for 240 seconds at a frequency of 2Hz, amplitude of 45 V and a tip temperature between 40 - 42°C. If the pain involved more than a single nerve root, the electrode was placed at a different segment and the procedure repeated. Outcome measures included the pain intensity score on a 0 - 10 numeric rating scale (NRS) and the Italian Pain Questionnaire (QUID) at pre-treatment, one and 6 months post-treatment. P values < 0.05 were considered statistically significant. RESULTS: In comparison with pre-treatment, a significant reduction in pain score was observed in mean NRS either at one and 6 months (P < 0.001). The QUID - Pain Rating Index rank displayed a parallel trend at the first (P < 0.001) and last follow-up (P = 0.01). Moreover, a direct correlation between the 2 scales occurred, showing a parallel score decreasing (P < 0.001). Eighteen (52.9%) and 17 (50%) of 34 patients showed pain reduction in NRS > 2 points and > 30%, at one and 6 months, respectively. LIMITATIONS: The non-controlled design of the study, the patients were heterogeneous, the small number of patients, and the duration of follow-up was limited to 6 months. CONCLUSIONS: PRF of dorsal root ganglion performed with a multifunctional electrode for > 240 seconds appears to be safe and might be more effective than the classic 120 seconds needle-mediated approach. Therefore, it may be considered as a valuable tool for the treatment of lumbosacral radicular pain with neuropathic features.