Intervertebral disc and endplate cells response to IL-1ß inflammatory cell priming and identification of molecular targets of tissue degeneration.

Inflammation represents an important factor leading to metabolic imbalance within the intervertebral disc (IVD), conducive to degenerative changes. Therefore, a thorough knowledge of the IVD and endplate (EP) cell behaviour in such pathological environments is essential when designing regenerative therapeutic strategies. The present study aimed at assessing the molecular response of the IVD constitutive nucleus pulposus (NPCs)-, annulus fibrosus (AFCs)- and endplate (EPCs)-derived cells to interleukin (IL)-1ß treatment, through large-scale, high-throughput microarray and protein analysis, identifying the differentially expressed genes and released proteins. Overall, the inflammatory stimulus downregulated stemness genes while upregulating pro-inflammatory, pro-angiogenic and catabolic genes, including matrix metalloproteases, which were not balanced by a concomitant upregulation of their inhibitors. Upregulation of anti-inflammatory and anabolic tumour necrosis factor inducible gene 6 protein (TNFAIP6), of IL-1 receptor antagonist (IL-1Ra) (at gene and protein levels) and of trophic insulin-like growth factor 1 (IGF1) was also observed in all cell types; IGF1 particularly in AFCs. An overall inhibitory effect of tumour necrosis factor alpha (TNFα) signal was observed in all cell types; however, EPCs showed the strongest anti-inflammatory behaviour. AFCs and EPCs shared the ability to limit the activation of the signalling mediated by specific chemokines. AFCs showed a slightly senescent attitude, with a downregulation of genes related to DNA repair or pro-mitosis. Results allowed for the identification of specific molecular targets in IVD and EP cells that respond to an inflammatory environment. Such targets can be either silenced (when pathological targets) or stimulated to counteract the inflammation.

Intervertebral disc and endplate cell characterisation highlights annulus fibrosus cells as the most promising for tissue-specific disc degeneration therapy.

Degenerative processes of the intervertebral disc (IVD) and cartilaginous endplate lead to chronic spine pathologies. Several studies speculated on the intrinsic regenerative capacity of degenerated IVD related to the presence of local mesenchymal progenitors. However, a complete characterisation of the resident IVD cell populations, particularly that isolated from the endplate, is lacking. The purpose of the present study was to characterise the gene expression profiles of human nucleus pulposus (NPCs), annulus fibrosus (AFCs) and endplate (EPCs) cells, setting the basis for future studies aimed at identifying the most promising cells for regenerative purposes. Cells isolated from NP, AF and EP were analysed after in vitro expansion for their stemness ability, immunophenotype and gene profiles by large-scale microarray analysis. The three cell populations shared a similar clonogenic, adipogenic and osteogenic potential, as well as an immunophenotype with a pattern resembling that of mesenchymal stem cells. NPCs maintained the greatest chondrogenic potential and shared with EPCs the loss of proliferation capability during expansion. The largest number of selectively highly expressed stemness, chondrogenic/tissue-specific and surface genes was found in AFCs, thus representing the most promising source of tissue-specific expanded cells for the treatment of IVD degeneration.

Acinic cell carcinoma of the parotid gland: from pathogenesis to management: a literature review.

PURPOSE: Acinic cell carcinoma (ACCs) is uncommon malignant epithelial neoplasm of the salivary glands; the most common presentation is a well-defined painless solid mass. Diagnosis of ACCs is frequently complicated, due to its similarity with benign tumors. METHODS: A review of the literature available on ACCs was carried out. Studies were sourced from PubMed with searching of relevant headings and sub-headings and cross-referencing. RESULTS: There are no clear characteristics of ACCs found on CT, MRI and ultrasound imaging. The management of the ACC, a rare malignancy of the parotid gland, is often difficult and controversial. Radical surgery is the best treatment option. The role of radiotherapy remains controversial: the precise indications and oncologic effects of adjuvant radiotherapy in ACC of the parotid gland are not well known. There is insufficient literature regarding the chemotherapy for metastatic ACC. CONCLUSION: Knowledge about ACC, a rare malignancy of parotid gland, has changed over the past few decades. More clinical randomized works would be needed, both to assess the real effectiveness of radio and chemotherapy and to have an unanimous consensus about their indications.

Multiresistant Acinetobacter baumannii causing malignant otitis externa: a case report.

Abstract: Malignant otitis externa is an infection of the skin and soft tissue of the ear canal, spreading to the nearby structures. It causes severe otalgia and otorrhea, and can lead to ominous consequences such as cranial nerve damage and meningitis. The main etiologic agent is Pseudomonas aeruginosa and treatment relies on broad-spectrum intravenous antibiotics. We report a rare case of a woman suffering from Malignant otitis externa caused by Acinetobacter baumannii and requiring the use of colistin.

Multifunctional CD4⁺ T cells in patients with American cutaneous leishmaniasis.

Leishmaniasis is a group of important parasitic diseases affecting millions worldwide. To understand more clearly the quality of T helper type 1 (Th1) response stimulated after Leishmania infection, we applied a multiparametric flow cytometry protocol to evaluate multifunctional T cells induced by crude antigen extracts obtained from promastigotes of Leishmania braziliensis (LbAg) and Leishmania amazonensis (LaAg) in peripheral blood mononuclear cells from healed cutaneous leishmaniasis patients. Although no significant difference was detected in the percentage of total interferon (IFN)-γ-producing CD4(+) T cells induced by both antigens, multiparametric flow cytometry analysis revealed clear differences in the quality of Th1 responses. LbAg induced an important proportion of multifunctional CD4(+) T cells (28% of the total Th1 response evaluated), whereas LaAg induced predominantly single-positive cells (68%), and 57% of those were IFN-γ single-positives. Multifunctional CD4(+) T cells showed the highest mean fluorescence intensity (MFI) for the three Th1 cytokines assessed and MFIs for IFN-γ and interleukin-2 from those cells stimulated with LbAg were significantly higher than those obtained after LaAg stimulation. These major differences observed in the generation of multifunctional CD4(+) T cells suggest that the quality of the Th1 response induced by L. amazonensis antigens can be involved in the mechanisms responsible for the high susceptibility observed in L. amazonensis-infected individuals. Ultimately, our results call attention to the importance of studying a Th1 response regarding its quality, not just its magnitude, and indicate that this kind of evaluation might help understanding of the complex and diverse immunopathogenesis of American tegumentary leishmaniasis.

Hyperosmia after COVID-19: hedonic perception or hypersensitivity?

OBJECTIVE: COVID-19 has been associated with a wide range of quantitative and qualitative disorders of smell, including hyposmia/anosmia, parosmia, and phantosmia; however, no reports to date have reported hyperosmia as a sequela of SARS-CoV-2 infection. PATIENTS AND METHODS: We present two cases of subjective hyperosmia in a South Tyrolean Alps family, occurring within days after recovery from SARS-CoV-2 infection with transient anosmia. RESULTS: The subjects, a mother and son, exhibited subjective hyperosmia despite normal objective olfactory testing. During independent assessments, the severity of hyperosmia and specific odors affected were highly correlated, consistent with shared genetic and environmental factors. In contrast, two other family members with COVID-19 had no perceptual distortion and normal recovery of smell. CONCLUSIONS: Subjective hyperosmia after COVID-19 infection exhibited striking similarity in two affected family members, suggesting interaction of environment, genetics, and perception.

Nasal Vestibulitis and Vestibular Furunculosis: a systematic review about two common nasal infections and considerations about correct diagnosis and management.

Background: Nasal vestibulitis (NV) and nasal vestibular furunculosis (NVF) are two infectious processes of the nasal vestibule, sharing common etiology, the same risk of complications, and similar treatment while remaining two different pathological entities. Methods: We performed a comprehensive literature research on NV and NVF in PubMed, Cochrane, and Google Scholar databases, with the aim to review the evidence on these two conditions and discuss the therapeutic approaches. Results: We identified a total of 248 records; according to our inclusion/exclusion criteria, 27 of them, published over a period of 59 years (1962-2021), were included in this review. Conclusion: NV and NVF are reported to be common conditions, with well-known etiological agents and risk factors. The diagnosis is clinical and topical antibiotics are the mainstay of treatment. Complications appear to be infrequent. Further studies are necessary to clarify the pathogenetic mechanisms and the exact prevalence of both conditions.

Evaluation of Discomfort in Nasopharyngeal Swab Specimen Collection for SARS-CoV-2 Diagnosis.

BACKGROUND: The rapid spread of COVID-19 worldwide has impo-sed the need to identify a test that quickly recognizes affected subjects, both symptomatic and asymptomatic. The most reliable option has been proven to be the RT-PCR, which allows to detect virus RNA on a specimen from a high viral load site, such as nasopharynx. Nasopha-ryngeal sample collection is possible by means of a nasopharyngeal swab (NPS) and is a practical and relatively non-invasive technique, but rather bothersome for the recipient. AIM: The aim of the present study is to evaluate the discomfort evoked during NPS. MATERIALS AND METHODS: We surveyed 429 patients receiving NPS before hospitalization or other procedures non related to COVID-19. For each one we noted the discomfort level felt during the swab using a 11-point numeric rating scale (NRS) for pain and the total time needed for the procedure to be taken. Sex, age, smoking status and positive history of previous swab have been taken into account. RESULTS: We found that, among the variables, sex had a statistically significant impact on the perceived discomfort of nasal swab, with females experiencing slightly more discomfort. CONCLUSIONS: NPS is largely a none-to-minimum discomfort in-ducing procedure. The differences in perceived discomfort could be explained based on anatomical features, and should remark the need for a tailored and anatomy-oriented approach in each patient.

Meningoencephalitis Associated with SARS-Coronavirus-2.

The aim of this work is to clarify the incidence of meningitis/encephalitis in SARS-CoV-2 patients. We conducted an initial search in PubMed using the Medical Subject Headings (MeSH) terms "meningitis," and "encephalitis,", and "COVID-19" to affirm the need for a review on the topic of the relationship between meningitis/encephalitis and SARS-CoV-2 infection. We included case series, case reports and review articles of COVID-19 patients with these neurological symptoms. Through PubMed database we identified 110 records. After removal of duplicates, we screened 70 record, and 43 were excluded because they focused on different SARS-CoV-2 neurological complications. For eligibility, we assessed 27 full-text articles which met inclusion criteria. Seven articles were excluded, and twenty studies were included in the narrative review, in which encephalitis and/or meningitis case reports/case series were reported. Neurological manifestations of COVID-19 are not rare, especially meningoencephalitis; the hypoxic/metabolic changes produced by the inflammatory response against the virus cytokine storm can lead to encephalopathy, and the presence of comorbidities and other neurological diseases, such as Alzheimer's disease, predispose to these metabolic changes. Further study are needed to investigate the biological mechanisms of neurological complications of COVID-19.

Drug-Induced Sleep Endoscopy (DISE) with Simulation Bite to Predict the Success of Oral Appliance Therapy in Treating Obstructive Sleep Apnea/Hypopnea Syndrome (OSAHS).

STUDY OBJECTIVES: Oral appliances have gained their place in the treatment of obstructive sleep apnea (OSA) where custom-made titratable mandibular advancement devices (MAD) have become the oral appliance of choice. This study aimed to asses the value of the drug-induced sleep endoscopy (DISE) using a MAD in the prediction of treatment outcome for OSAHS. METHODS: This is a prospective, single-center cohort study that enrolled sixty-six consecutive patients with diagnosed OSA (5 events/h < apnea-hypopnea index (AHI) < 50 events/h) to be treated with a custom-made titratable MAD. The patients were evaluated polysomnographically with the MAD in situ after the adaptation and titration period of 3 months. The associations between findings during DISE and treatment outcome were assessed. RESULTS: The subjects showed a wide range of severity of OSAHS pre-treatment: median AHI was 43.10 with a range from 20.13 to 66.07. The simulation bite was associated with a significant increase in cross-sectional area at level of the velopharynx, tongue base and epiglottis. MAD treatment response in the studied population was 91%, with a mean AHI improving from 43.10 to 12.93. CONCLUSIONS: Drug-induced sleep endoscopy with simulation bite is an acceptably reproducible technique for determining the sites of obstruction in OSAHS subjects; it thus offers possibilities as a prognostic indicator for treatment with MAD.

Dietary Restriction for The Treatment of Meniere's Disease.

Meniere's disease (MD) is an idiopathic inner ear disorder characterized by spontaneous recurrent vertigo, fluctuating sensorineural hearing loss (SNHL), aural fullness and tinnitus. Endolymphatic hydrops (EH) of the inner ear is currently considered the pathophysiological mechanisms that underlies typical symptoms of MD. MD diagnosis is based on the criteria of the Baràny Society. There are many therapeutic options for MD, but none is considered effective by the scientific community. The first-line treatment commonly includes dietary modification, as low salt diet and reduction of alcohol and caffeine daily intake. Although some studies showed a positive effect of these dietary restrictions, even in the prevention of recurrences, currently there is no uniform consensus on their usefulness. New dietary approach, such SPC-flakes, are being evaluated: further assessments will be needed to validate their use in clinical practice.

Sub-Clinical Effects of Chronic Noise Exposure on Vestibular System.

AIM: to investigate the effect of chronic noise exposure on vestibular function of subjects without clinical evidence of vestibular disorders and with documented cochlear damage from noise. SUBJECTS AND METHODS: 25 patients with chronic noise-induced hearing loss (NIHL) and without vestibular complaints (group A) and 25 matched controls with sensorineural hearing loss without noise exposure (group B), underwent audiological and vestibular test including caloric and cervical vestibular-evoked myogenic potentials tests (cVEMPs). RESULTS: In subjects chronically exposed to noise, similarly to that of the auditory threshold, an increase in the evocation threshold of VEMPs has been documented, statistically significant (p<0,05) and independent of the performance of the auditory threshold. p1-n1 amplitude values showed a significant difference between group A and group B. No significant difference for p1-n1 latencies between the two groups was found. CONCLUSION: We have documented the possibility of vestibular lesion, along with cochlear damage, related to chronic acoustic trauma.

No evidence of major transcriptional changes in the brain of mice exposed to 1800 MHz GSM signal.

To analyze possible effects of microwaves on gene expression, mice were exposed to global system for mobile communication (GSM) 1800 MHz signal for 1 h at a whole body SAR of 1.1 W/kg. Gene expression was studied in the whole brain, where the average SAR was 0.2 W/kg, by expression microarrays containing over 22,600 probe sets. Comparison of data from sham and exposed animals showed no significant difference in gene expression modulation. However, when less stringent constraints were adopted to analyze microarray results, 75 genes were found to be modulated following exposure. Forty-two probes showed fold changes ranging from 1.5 to 2.8, whereas 33 were down-regulated from 0.67- to 0.29-fold changes, but these differences in gene expression were not confirmed by real-time PCR. Under these specific limited conditions, no consistent indication of gene expression modulation in whole mouse brain was found associated to GSM 1800 MHz exposure.

Class A helix-loop-helix proteins are positive regulators of several cyclin-dependent kinase inhibitors' promoter activity and negatively affect cell growth.

The class A of basic helix-loop-helix (bHLH) proteins are ubiquitously expressed transcription factors playing a pivotal role in the regulation of cell growth and differentiation. We determined that enforced expression of all four different mammalian members of this family, E12, E47, E2-2, and HEB, suppresses the cell colony-forming efficiency of several cell lines. To gain insights into the mechanisms by which class A bHLH factors affect cell growth, we have investigated their role in the transcriptional regulation of cyclin-dependent kinase inhibitors. We found that p21CIP1/ WAF1, p15INK4B, and p16INK4B promoter sequences contain E-boxes that render these genes competent for class A bHLH-mediated transcriptional activation and Id-mediated repression. The mechanism underlying the class A bHLH-mediated inhibition of cell growth does not involve an arrest of G1 progression in 293T cells. In fact, contrary to what has been found in 3T3 NIH fibroblasts, we found that enhanced expression of class A bHLH proteins led to a decreased proliferation rate by promoting cell death associated with the induction of apoptosis. These findings highlight the role of the class A bHLH proteins as general negative regulators of cell proliferation through a mechanism(s) that involves both enhancement of several cyclin-dependent kinase inhibitor genes expression and promotion of cell death.

Differential transcriptional regulation of c-myc promoter through the same DNA binding sites targeted by Sp1-like proteins.

Sp1 and Sp3 are closely related members of a gene family encoding proteins with very similar structural features. The zinc finger DNA binding domains of Sp1 and Sp3 are highly conserved and they bind to GC and GT box with comparable affinities. To begin to delineate the specific roles of these two members of the Sp1-like gene family, here we have analysed the DNA binding specificity and their effects on activation of human c-myc promoter. We found that both proteins bind to the same sites of c-myc promoter, upstream to both the P1 and P2 initiation sites. Cotransfection experiments, in mammalian and insect cells, indicated that Sp1 trans-activate c-myc promoter, whereas Sp3 did not. In addition, enforced expression of Sp3 repressed Sp1-mediated activation of c-myc. Finally, we found that Sp1 and E2F-1/DP-1 cooperatively trans-activate c-myc promoter. In contrast enforced expression of Sp3 fails to repress E2F-1/DP-1-mediated activation.

B-MYB transactivates its own promoter through SP1-binding sites.

B-MYB is an ubiquitous protein required for mammalian cell growth. In this report we show that B-MYB transactivates its own promoter through a 120 bp segment proximal to the transcription start site. The B-MYB-responsive element does not contain myb-binding sites and gel-shift analysis shows that SP1, but not B-MYB, protein contained in SAOS2 cell extracts binds to the 120 bp B-myb promoter fragment. B-MYB-dependent transactivation is cooperatively increased in the presence of SP1, but not SP3 overexpression. When the SP1 elements of the B-myb promoter are transferred in front of a heterologous promoter, an increased response to B-MYB results. In contrast, c-MYB, the prototype member of the Myb family, is not able to activate the luciferase construct containing the SP1 elements. With the use of an SP1-GAL4 fusion protein, we have determined that the cooperative activation occurs through the domain A of SP1. These observations suggest that B-MYB functions as a coactivator of SP1, and that diverse combinations of myb and SP1 sites may dictate the responsiveness of myb-target genes to the various members of the myb family.

Purification and partial characterization of single-chain ribosome-inactivating proteins from the seeds of Phytolacca dioica L.

Three ribosome-inactivating proteins (RIPs) similar to those already known (Stirpe et al. (1992) Bio/Technology 10, 405-412) were purified from the seeds of Phytolacca dioica. These proteins, called Phytolacca dioica RIPs (PD-S1, PD-S2 and PD-S3 RIPs), are glycoproteins, with M(r) approx. 30,000, inhibit protein synthesis by a rabbit reticulocyte lysate and phenylalanine polymerization by isolated ribosomes, and depurinate rat liver rRNA in an apparently identical manner as the A-chain of ricin and other RIPs (Endo et al. (1987) J. Biol. Chem. 262, 5908-5912). Part of the purified rat liver ribosomes appeared resistant to the action of PD-S RIPs. The most abundant protein, PD-S2 RIP, gave a weak or nil cross-reaction with sera against various other RIPs, including a pokeweed antiviral protein from the roots of Phytolacca americana. PD-S2 RIP was linked to a monoclonal antibody (Ber-H2) against the CD30 human lymphocyte antigen and the resulting immunotoxin was selectively toxic to the CD30 + Hodgkin's lymphoma-derived L540 cell line.

In vitro T-cell profile induced by BCG Moreau in healthy Brazilian volunteers.

Tuberculosis (TB) remains the world's leading cause of morbidity and mortality. Although Mycobacterium bovis bacillus Calmette-Guérin (BCG) is the only vaccine currently in use, its efficacy is highly variable. It has been suggested that early antigenic presentation is a pivotal event leading to a better immune response in TB vaccine models. To investigate this further, we compared in vitro cell-mediated immune responses in the context of early sensitization with TB (i.e. healthy adults vaccinated with BCG when they were young, HD; n = 25) to those in its absence (i.e., newborns with naïve immunity to TB, UV; n = 10) by challenging mononuclear cells with BCG Moreau. After 48 hours, CD4+ and CD8+ T cells were harvested from both groups and stained for PD-1/CD25/ FOXP3. In addition, supernatants were assayed for a broad range of cytokines using an array system. The HD group showed robust reactivity to Protein Purified Derivative and BCG while the naïve, UV group did not. Similarly, in terms of PD-1 expression and Treg cells (CD4+/CD25high(+)/FOXP3+), only the HD group showed higher levels in CD4 lymphocytes. Otherwise, only the UV group showed expression of CD25dim+ as an activation marker dependent on BCG infection. In terms of cytokines, the HD group showed higher levels of Th1 (IL-2/TNF-α/IFN-γ) and regulatory (IL-10) profiles, with monocytes, but not Tr1 cells, acting as the main source of IL-10. Taken together, our results highlight critical roles of early sensitization with TB in mounting cell-mediated immune responses.

Transcriptional regulation by the Sp family proteins.

Sp1 is one of the very first cellular transcription factors to be identified and cloned in virtue of its binding to a G-rich motif in the SV40 early promoter. Sp1 protein binds to the G-rich sequences present in a variety of cellular and viral promoters and stimulates their transcriptional activity. Recently, a number of other GC and/or GT box-binding factors homologous to Sp1 have been isolated, namely Sp2, Sp3 and Sp4, and the two more distantly related factors, BTEB and BTEB2. The discovery of this family highlights a previously unknown level of complexity of transcriptional regulation of promoters containing GC and/or GT box motifs. This review focuses primarily on strategies aimed to elucidate the transcription properties of the Sp1-like factors and discusses the experimental problems inherent in the attempt to define their respective functions.