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Could it have been the remoteness in which I now find myself, or the peculiarity of this house, of this road, of the name of this road, to impose on me these nocturnal wanderings of the mind which, for some time now, I have become used to living with? Who or what is the true author of this writing, I will not be able to determine, and if at the beginning I could still venture the illusion of autonomous choices, it has been enough, after having gone through a few dozen pages, for me to realize that, slowly or in a rapid hurry, I can no longer guarantee to maintain a firm control of the contents. As to the late professor of latinitas, inhabitant of the floors of this house, which now I, in his place, am treading, I would never have imagined him capable of opening up to me such a necessary perlustration into the protagonists and the anxieties of my life. (read more)

Age-related macular degeneration (AMD) and related macular dystrophies (MDs) primarily affect the retinal pigment epithelium (RPE) in the eye. A hallmark of AMD/MDs that drives later-stage pathologies is drusen. Drusen are sub-RPE lipid-protein-rich extracellular deposits, but how drusen forms and accumulates is not known. We utilized human induced pluripotent stem cell (iPSC)-derived RPE from patients with AMD and three distinct MDs to demonstrate that reduced activity of RPE-secreted matrix metalloproteinase 2 (MMP2) contributes to drusen in multiple maculopathies in a genotype-agnostic manner by instigating sterile inflammation and impaired lipid homeostasis via damage-associated molecular pattern molecule (DAMP)-mediated activation of receptor for advanced glycation end-products (RAGE) and increased secretory phospholipase 2-IIA (sPLA2-IIA) levels. Therapeutically, RPE-specific MMP2 supplementation, RAGE-antagonistic peptide, and a small molecule inhibitor of sPLA2-IIA ameliorated drusen accumulation in AMD/MD iPSC-RPE. Ultimately, this study defines a causal role of the MMP2-DAMP-RAGE-sPLA2-IIA axis in AMD/MDs., Competing Interests: Declaration of interests University of Rochester has filed a provisional US patent application: U.S. Provisional Patent Application No. 63/632,123, filed April 10, 2024, title: “Drug Treatment for Macular Degeneration.”, (Published by Elsevier Inc.)

Carbon dots (CDs) have attracted much attention due to their excellent properties and applications, especially the use for gene delivery. Considering the risks and concerns involved in the use of viral vectors for gene delivery in vivo, non-viral vectors such as CDs have gradually become an ideal alternative due to their biocompatibility and low toxicity. Therefore, in this study, the potential to apply CDs as a non-viral vector for gene delivery was investigated. The CDs were prepared using citric acid and pentaethylenehexamine (PEHA) as precursors via a one-step microwave-mediated approach. The optical, structural, and morphological properties of PEHA-derived CDs (PCDs) were characterized by ultra-violet spectroscopy (UV-vis), photoluminescence (PL), Fourier Transform Infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), X-ray photoelectron spectroscopy (XPS), zeta potential, circular dichroism spectrometry, atomic force (AFM) and transmission electron microscopies (TEM). The analysis demonstrated that the as-prepared PCDs were rich in amine groups and were positively charged. Subsequently, gel retardation assay showed that PCDs could non-covalently bind with DNA at a mass ratio of 2:1 (PCDs: DNA). Additionally, PCDs possessed a tremendously lower cytotoxicity compared with polyethylenimine (PEI), a popular precursor/dopant for many CDs preparations, and their plasmid composite showed a high transfection efficiency. Meanwhile, PCDs were also observed to cross the blood-brain barrier (BBB) by using a zebrafish model. In conclusion, these results significantly indicate that PCDs are a potential non-viral nucleic acid/gene vector to gene therapy. Also, PCDs can be utilized in drug delivery for treating brain diseases, such as Alzheimer's disease and brain tumors., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Nucleus targeting is tremendously important in cancer therapy. Cationic carbon dots (CCDs) are potential nanoparticles which might enter cells and penetrate nuclear membranes. Although some CCDs have been investigated in nucleus targeting and applied in nuclear imaging, the CCDs derived from drugs, that are able to target the nucleus, bind with DNA and inhibit the growth of cancer cells have not been reported. In this project, 1, 2, 4, 5-benzenetetramine (Y15, a focal adhesion kinase inhibitor) derived cationic carbon dots (Y15-CDs) were prepared via a hydrothermal approach utilizing Y15, folic acid and 1,2-ethylenediamine as precursors. Based on the structural, optical, and morphologic characterizations, Y15-CDs possess rich amine groups and nitrogen in structure, an excitation-dependent photoluminescence emission, and a small particle size of 2 to 4 nm. The DNA binding experiments conducted through agarose gel electrophoresis, UV-vis absorption, fluorescence emission, and circular dichroism spectroscopies, prove that Y15-CDs might bind with DNA via electrostatic interactions and partially intercalative binding modes. In addition, the cell imaging and cytotoxicity studies in human foreskin fibroblasts (HFF), prostate cancer (PC3) and osteosarcoma cells (U2OS) indicate the nucleus targeting and anticancer abilities of Y15-CDs. Most interestingly, Y15-CDs exhibit a higher cytotoxicity to cancer cells (PC3 and U2OS) than to normal cells (HFF), inferring that Y15-CDs might be potentially applied in cancer therapy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Objective: Placebo-induced adverse events, or nocebo effects, occur when doctor-patient communication anticipates the onset of negative symptoms. They have been found to correlate with the anxiety-related activity of the hypothalamic-pituitary-adrenal system. Here we try to determine if prenatal hyperactivity of this system, as assessed through plasma cortisol, may influence nocebo effects in adulthood., Method: We investigated the rate and magnitude of nocebo effects in 378 adults whose prenatal maternal plasma cortisol was measured during the first, second and third trimester of pregnancy. The healthy subjects underwent a nocebo oxygen challenge. This consisted of the inhalation of fake (placebo) oxygen and assessment of the following adverse events: headache, chest pain, abdominal pain, and cough. Plasma cortisol responses during the nocebo adverse events were also measured., Results: 41 out of 46 (89.1%) subjects who reported 3 adverse events, and 37 out of 37 (100%) subjects who reported 4 adverse events had prenatal maternal cortisol above normal levels. By contrast, only 10 out of 143 (7%) subjects who reported 0 adverse events showed prenatal maternal cortisol above the normal range. Moreover, whereas subjects who reported 3 and 4 adverse events showed a significant increase in plasma cortisol following the nocebo challenge, subjects who reported 0 adverse events showed no changes., Conclusions: These findings emphasize the importance of the doctor-patient communication in perceiving symptoms like pain, and suggest that those subjects with high prenatal maternal cortisol may be more sensitive to the effects of a negative communication in adulthood., Competing Interests: Competing interests: None., (© 2022 Giovanni Fioriti Editore s.r.l.)

Cerebral Cavernous Malformation (CCM) is a cerebrovascular disease of genetic origin that predisposes to seizures, focal neurological deficits and fatal intracerebral hemorrhage. It may occur sporadically or in familial forms, segregating as an autosomal dominant condition with incomplete penetrance and highly variable expressivity. Its pathogenesis has been associated with loss-of-function mutations in three genes, namely KRIT1 (CCM1), CCM2 and PDCD10 (CCM3), which are implicated in defense mechanisms against oxidative stress and inflammation. Herein, we screened 21 Italian CCM cases using clinical exome sequencing and found six cases (~29%) with pathogenic variants in CCM genes, including a large 145−256 kb genomic deletion spanning the KRIT1 gene and flanking regions, and the KRIT1 c.1664C>T variant, which we demonstrated to activate a donor splice site in exon 16. The segregation of this cryptic splicing mutation was studied in a large Italian family (five affected and seven unaffected cases), and showed a largely heterogeneous clinical presentation, suggesting the implication of genetic modifiers. Moreover, by analyzing ad hoc gene panels, including a virtual panel of 23 cerebrovascular disease-related genes (Cerebro panel), we found two variants in NOTCH3 and PTEN genes, which could contribute to the abnormal oxidative stress and inflammatory responses to date implicated in CCM disease pathogenesis.

Vascular malformations include various disorders characterized by morphological, structural and/or functional alterations of blood and lymph vessels. Most are sporadic, due to somatic mutations. Here, we report a cohort of patients with sporadic and/or unifocal vascular malformations, in whom we carried out next generation sequencing analysis of a panel of genes associated with vascular malformations. The 115 patients analyzed were from different clinical centres. In 37 patients (32%), we found pathogenic mutations: most of these were gain-of-function mutations in PIK3CA (18%, 21/115) and TEK (13/115, 11%). We also found mutations in GNAQ , CCM2 and PTEN . Identifying pathogenic variants in patients with vascular malformations can help improve management, particularly in cases with activating mutations that cause an increase in cell proliferation. Personalized pharmacological treatment, if possible, is now considered preferable to surgery and can help prevent recurrences, i.e., long-term complications of residual malformation or regrowth of tumors. For instance, rapamycin is currently being investigated for the treatment of various vascular malformations associated with hyperactivation of the phosphoinositide 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway.

Background: Smith Lemli Opitz syndrome (SLOS; OMIM #270400) is an autosomal recessive metabolic disorder caused by mutations in the DHCR7 gene. SLOS is characterized by a plethora of abnormalities involving mainly the brain and the genitalia but also the cardiac, skeletal and gastroenteric system, typical dysmorphic facial features, and variable degrees of developmental delay and intellectual disability (ID). SLOS has a broad phenotypic spectrum, ranging from multiple congenital malformation syndrome, to mild developmental delay and minor malformations. A large number of mutations have been described in the DHCR7 gene, with few common mutations accounting for the majority of mutated alleles found in patients and a large number of very rare or even private variants. Due to the wide variety of clinical presentations, diagnosis can be difficult, especially in the milder forms of the disorder. Furthermore, establishing a molecular diagnosis can be complicated by finding variants of unknown clinical significance in such cases., Case Presentation: We report a case of SLOS at the mild end of the clinical spectrum, presenting with bilateral pelvis ectasia, mild dysmorphic features and mild intellectual disability. The case is compound heterozygous for a known pathogenic mutation (c.724C > T, p.Arg242Cys) and a mutation that has only been reported once in a Portuguese patient (c.521 T > C, p.Phe174Ser) whose pathogenicity has not been yet assessed. We compared the two patients carrying the p.Phe174Ser variant and concluded that this variant is associated with mild forms of SLOS., Conclusion: We report a patient with a mild case of SLOS, highlighting the importance of recognizing subtle anomalies of the genitourinary system, associated with mild dysmorphic features and mild intellectual disability in establishing the diagnosis of mild forms of SLOS. With this report, we confirm the pathogenicity of the p.Phe174Ser variant and we also provide evidence of its association with mild forms of SLOS. This finding further facilitates the establishment of a genotype-phenotype correlation for SLOS. This helps in counselling for this disorder and in predicting therapeutic responses.

Purpose: PAX6 mutations cause aniridia as well as other various congenital eye abnormalities. Aniridia can be due to both point mutations and chromosomal deletions/rearrangements. Therefore, a complete search for PAX6 gene alterations in aniridia subjects requires a technically complex approach involving the comprehension of fluorescence in situ hybridization (FISH) analysis. In the present study, an Italian casistic of aniridia patients has been investigated and a quantitative polymerase chain reaction (PCR) assay to detect PAX6 gene deletions was set up., Methods: Twenty-one aniridia patients were screened for point mutations (missense, nonsense, splicing-affecting, and short insertion/deletion) by using single-stranded conformational polymorphism (SSCP) and denaturing high performance liquid chromatography (dHPLC). To reveal deletions not detectable by SSCP or dHPLC, a quantitative PCR approach was set up for the PAX6 structural gene and for regions 5' and 3' to it at the level of WT1 and ELP4, respectively., Results: Point mutations were found in 7 out of 21 patients. Three out of twenty-one patients showed deletions at the level of the PAX6 structural gene. In addition, two familial cases showed an undamaged PAX6 gene but a deletion in the region 3' to it at level of the ELP4 gene. In one of the families, the presence of the deletion has been confirmed by linkage analysis of polymorphic markers., Conclusions: In our casistic, a significant fraction of familial aniridia patients appears to be caused by a 3' deletion to PAX6, suggesting that evaluation of this alteration should be included in routine procedures of aniridia patients analysis. The quantitative PCR assay described here represents a simple approach to accomplish this task.

Several diagnostic strategies have been applied to the detection of FMR1 gene repeat expansions in fragile X syndrome. Here, we report a novel polymerase chain reaction-based strategy using the Expand Long Template PCR System (Roche Diagnostics, Mannheim, Germany) and the osmolyte betaine. Repeat expansions up to approximately 330 CGGs in males and up to at least approximately 160 CGGs in carrier women could be easily visualized on ethidium bromide agarose gels. We also demonstrated that fluorescence analysis of polymerase chain reaction products was a reliable tool to verify the presence of premutation and full mutation alleles both in males and in females. This technique, primarily designed to detect premutation alleles, can be used as a routine first screen for expanded FMR1 alleles.

It has been suggested that genes other than CFTR could modulate the severity of lung disease in cystic fibrosis (CF). Neutrophil Fcgamma receptor II (FcgammaRII) is involved in host defense against microorganisms and in inflammatory response. We evaluated the association between genetic variability of this gene and both airway infection with Pseudomonas aeruginosa and severity of lung disease in patients with CF. We studied 167 Italian unrelated patients with CF and 50 control subjects. The distribution of FcgammaRIIA genotypes in CF patients was compared with that in control subjects and the different genotypes were related with the presence or absence of P. aeruginosa infection and markers of disease severity in CF patients. The distribution of FcgammaRIIA genotypes was not significantly different between CF patients and controls. We observed that in CF patients with the same CFTR genotype (DeltaF508/DeltaF508), those carrying the R allele of FcgammaRIIA had an increased risk of acquiring chronic P. aeruginosa infection (P=0.042, R.R.: 4.38; 95% CI: 1.17/22.4). Moreover, the frequency of R/R genotype in patients with chronic P. aeruginosa infection seems to be higher than that of control subjects and patients without chronic infection. The observation that CF patients carrying the R allele of FcgammaRIIA are at higher risk of acquiring chronic P. aeruginosa infection suggests that the FcgammaRII loci genetic variation is contributing to this infection susceptibility.

We induced specific expectations of analgesia on four different parts of the body to understand how endogenous opioid systems are activated by expectancies. The left hand, right hand, left foot, and right foot were simultaneously stimulated by means of a subcutaneous injection of capsaicin, which produces a painful burning sensation. Specific expectations of analgesia were induced by applying a placebo cream on one of these body parts and by telling the subjects that it was a powerful local anesthetic. In such a way, expectancy of the anesthetic effect was directed only toward the part on which the placebo cream was applied. We found that a placebo analgesic response occurred only on the treated part, whereas no variation in pain sensitivity was found on the untreated parts. If the same experiment was performed after an intravenous infusion of the opioid antagonist naloxone, this highly spatial-specific placebo response was totally abolished, indicating that it was completely mediated by endogenous opioid systems. These findings show that a spatially directed expectation of pain reduction is capable of inducing a specific effect only on the part of the body which is the target of the expectation. Most important, this specific effect is mediated by endogenous opioids, indicating that placebo-activated opioids do not act on the entire body but only on the part where expectancy is directed. This suggests that a highly organized and somatotopic network of endogenous opioids links expectation, attention, and body schema.