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Objectives: The presence of prostate‐specific antigen (PSA) in saliva and salivary glands has been reported. Nevertheless, its release pathway in these glands remains to be elucidated. Here, we showed PSA subcellular distribution focusing on its plausible route in human salivary parenchyma. Materials and Methods: Sections of parotid and submandibular glands were subjected to the immunohistochemical demonstration of PSA by the streptavidin–biotin method revealed by alkaline phosphatase. Moreover, ultrathin sections were collected on nickel grids and processed for immunocytochemical analysis, to visualize the intracellular distribution pattern of PSA through the observation by transmission electron microscopy. Results: By immunohistochemistry, in both parotid and submandibular glands PSA expression was detected in serous secretory acini and striated ducts. By immunocytochemistry, immunoreactivity was retrieved in the cytoplasmic compartment of acinar and ductal cells, often associated with small cytoplasmic vesicles. PSA labeling appeared also on rough endoplasmic reticulum and in the acini's lumen. A negligible PSA labeling appeared in most of the secretory granules of both glands. Conclusions: Our findings clearly support that human parotid and submandibular glands are involved in PSA secretion. Moreover, based on the immunoreactivity pattern, its release in oral cavity would probably occur by minor regulated secretory or constitutive‐like secretory pathways. [ABSTRACT FROM AUTHOR]

Abstract Background Cervical cancer ranks as the first most frequent cancer among women in Benin. The major cause of cervical cancer now recognized is persistent infection of Human Papillomavirus (HPV). In Benin there is a lack of screening programs for prevention of cervical cancer and little information exists regarding HPV genotype distribution. Methods Cervical cells from 725 women were examined for the presence of viral DNA by means of a polymerase chain reaction (PCR) multiplex-based assay with the amplification of a fragment of L1 region and of E6/E7 region of the HPV genome, and of abnormal cytology by Papanicolaou method. The association between HPV status and Pap test reports was evaluated. Socio-demographic and reproductive characteristics were also related. Results A total of 18 different HPV types were identified, with a prevalence of 33.2% overall, and 52% and 26.7% among women with and without cervical lesions, respectively. Multiple HPV infections were observed in 40.2% of HPV-infected women. In the HPV-testing group, the odds ratio for the detection of abnormal cytology was 2.98 (95% CI, 1.83-4.84) for HPV positive in comparison to HPV negative women. High risk types were involved in 88% of infections, most notably HPV-59, HPV-35, HPV-16, HPV-18, HPV-58 and HPV-45. In multiple infections of women with cytological abnormalities HPV-45 predominated. Conclusions This study provides the first estimates of the prevalence of HPV and type-specific distribution among women from Benin and demonstrates that the epidemiology of HPV infection in Benin is different from that of other world regions. Specific area vaccinations may be needed to prevent cervical cancer and the other HPV-related diseases.

Objective: The purpose of the study was to examine whether the insertion (I) and/or deletion (D) polymorphism of ACE confers susceptibility to primary pterygium in Sardinian patients in a case-control study. Methods and results: Polymorphism genotyping was performed by nested PCR using genomic DNA extracted from the whole peripheral blood of participants with (n=251) and without (n=260) pterygium. DD, ID and II genotype frequencies were: 48%, 39% and 13%, respectively, for patients with pterygium, and 15%, 40% and 44%, respectively, for the control group. A statistically significant difference was found between the pterygium and control groups for the ACE I/D polymorphism (p<0.001). Moreover, a statistically significant difference was found between the DD and II groups (p<0.01; OR=10.49; 95% CI 6.18 to 17.79), DD+ID versus II group (p<0.01; OR=5.23; 95% CI 3.37 to 8.13) and DD versus ID groups (p<0.01; OR=3.21; 95% CI 2.04 to 5.04). Conclusions: Statistical analysis showed that the DD genotype is associated with an increased risk of developing pterygium, and with a good chance that the D allele may play an important role in the development of disease. [ABSTRACT FROM AUTHOR]

Histological analysis allows an accurate classification of most melanocytic lesions as benign or malig-nant. However, a challenging diagnosis can be faced when differentiating a nevus from a melanoma, mostly due to the heterogeneous histological appearance of melanomas. It is thus necessary to use immunohistochemistry as a complementary tool. The immunohistochemical expression of tyrosinase, the key melanogenic enzyme in melanocytes, has often been useful in formulating a differential diagnosis thanks to the peculiar staining pattern in nevocytes compared with melanoma cells. The expression pattern of tyrosinase in nevi appears to parallel the cytoarchitectural changes typically observable within the lesion: nevus cells in the epidermis or in the superficial dermis are more likely to be larger and strongly express melanocytic differentiation antigens, such as tyrosinase, compared with deeper nevocytes [1]. Our study aimed to evaluate the immunohistochemical expression pattern of the tyrosinase antigen (clone T311) in different histological types of acquired dysplastic melanocytic nevi, including junctional, compound, and intradermal nevi, as well as in a panel of normal skin tissues. Moreover, to evaluate whether the expression of tyrosinase by nevus cells, pointing out the acquisition of melanin-producing capabilities, was associated with the differentiation state of nevocytes, we likewise investigated the immunohistochemical expression of the two markers of pluripotency, CD34 and nestin. Our results revealed a prominent immunoreactivity for tyrosinase in junctional and superficial dermal nevocytes and a decreasing gradient of staining in dermal nevocytes, up to become negative in the deeper dermis. All junctional and dermal nevocytes were negative for CD34. Nestin immunostaining showed an opposing pattern compared with tyrosinase, leading us to look into the melanocytic nevus as a "histopathological model" to trace the final stages of the differentiation pathway that neural crest-derived melanocyte precursors undertake toward their ultimate anatomical and functional site into the epidermis, consistently with Cramer's dermal precursor model of melanocytic origin and the stem cell-based concept of nevogenesis [2]. Both CD34 and nestin intensely stained the small aggregates of cells, resembling "niches", adjacent to the bulge area of hair follicles, that may represent the reservoir of neural crest-derived melanocyte stem cells residing in the dermis. [ABSTRACT FROM AUTHOR]

Telocytes (TCs) are a novel and peculiar interstitial cell type already described in many tissues and organs. Their name derives from their typical extremely long, thin, tortuous, and overlapping processes called telopodes (Tps), forming a stromal threedimensional network. TCs occupy a strategic position in relation to stem cell niches, blood capillaries, and nerve bundles, then contributing to maintain tissues homeostasis. However, TCs involvement in the pathophysiology of several disorders is being increasingly investigated because of their role as “connecting cells", mostly oriented to intercellular signalling. Previous study provided evidences for TCs involvement in neoangiogenesis (2), and we recently demonstrated their presence also in pterygium, a common degenerative and hyperplastic disorder of bulbar conjunctiva, characterized by an intense process of neovascularization. TCs and TPs were detected both in the subepithelial layer and in the connectival stroma of pterygium, especially in close relationship to the newly formed vessels. Since it is well established that TCs share the same ultrastructural features but display totally different morphology and immunophenotype based on their organ and tissue localization, the purpose of the study was to perform in pterygium a morphological and immunohistochemical analysis by light microscopy of thin and semithin sections and an ultrastructural study by transmission electron microscopy (TEM). The results will be discussed. [ABSTRACT FROM AUTHOR]

Background: One of the most challenging tasks of modern biology concerns the real-time tracking and quantification of mRNA expression in living cells. On this matter, a novel platform called SmartFlare TM has taken advantage of fluorophore-linked nanoconstructs for targeting RNA transcripts. Although fluorescence emission does not account for the spatial mRNA distribution, NanoFlare technology has grown a range of theranostic applications starting from detecting biomarkers related to diseases, such as cancer, neurodegenerative pathologies or embryonic developmental disorders., Aim: To investigate the potential of SmartFlare TM in determining time-dependent mRNA expression of prominin 1 ( CD133 ) and octamer-binding transcription factor 4 ( OCT4 ) in single living cells through differentiation., Methods: Brain fragments from the striatum of aborted human fetuses aged 8 wk postconception were processed to obtain neurospheres. For the in vitro differentiation, neurospheres were gently dissociated with Accutase solution. Single cells were resuspended in a basic medium enriched with fetal bovine serum, plated on poly-L-lysine-coated glass coverslips, and grown in a lapse of time from 1 to 4 wk. Live cell mRNA detection was performed using SmartFlare TM probes (CD133, Oct4, Actin, and Scramble). All the samples were incubated at 37 °C for 24 h. For nuclear staining, Hoechst 33342 was added. SmartFlare TM CD133- and OCT4-specific fluorescence signal was assessed using a semiquantitative visual approach, taking into account the fluorescence intensity and the number of labeled cells., Results: In agreement with previous PCR experiments, a unique expression trend was observed for CD133 and OCT4 genes until 7 d in vitro (DIV). Fluorescence resulted in a mixture of diffuse cytoplasmic and spotted-like pattern, also detectable in the contacting neural branches. From 15 to 30 DIV, only few cells showed a scattered fluorescent pattern, in line with the differentiation progression and coherent with mRNA downregulation of these stemness-related genes., Conclusion: SmartFlare TM appears to be a reliable, easy-to-handle tool for investigating CD133 and OCT4 expression in a neural stem cell model, preserving cell biological properties in anticipation of downstream experiments., Competing Interests: Conflict-of-interest statement: The authors declare that they have no conflict of interest., (©The Author(s) 2021. Published by Baishideng Publishing Group Inc. All rights reserved.)

Purpose: Telocytes (TCs) are peculiar interstitial cells, characterized by their typical elongated and interconnected processes called telopodes. TCs are supposed to contribute to maintain tissue homeostasis but also to be involved in the pathophysiology of many disorders. The aim of the study was to identify TCs in pterygium, a chronic condition of bulbar conjunctiva, and to examine possible differences in TCs in terms of immunophenotype and/or localization between pterygium and normal conjunctiva, to evaluate the possible involvement of TCs in pathogenesis of pterygium., Methods: The analysis of the immunophenotype of TCs was performed on a group of 40 formalin-fixed and paraffin-embedded primary pterygium and ten bulbar conjunctiva samples. We examined with immunohistochemistry the expression of 11 commercially available antibodies (PDGFRα, CD34, c-kit, nestin, vimentin, α-SMA, laminin, S100, VEGF, CD133, and CD31) and with double immunofluorescence the concomitant expression of PDGFRα and CD34, and PDGFRα and nestin. In addition, we performed an ultrastructural study with transmission electron microscopy (TEM) on a group of five pterygium and three conjunctiva biopsy specimens., Results: TCs, ultrastructurally identified according to their "moniliform" prolongations, were localized underneath the epithelium along the basement membrane, around the vessels, and near the nerves and scattered in the stroma. In contrast, TCs, as fibroblasts, were almost absent in the fibrotic areas. In pterygium and normal conjunctiva, the TCs shared the same distribution pattern, except a marked TC hyperplasia detected in pterygium. Moreover, in pterygium, the immunohistochemical analysis of TCs showed a strong immunoreactivity to PDGFRα, CD34, and nestin. This result was confirmed with double immunofluorescence labeling, revealing that in pterygium stromal TCs always showed a PDGFRα+/nestin+ and PDGFRα+/CD34+ immunophenotype. Furthermore, moderate staining to vimentin and VEGF was detected, but only a small number of cells were weakly immunoreactive to laminin and S100. Only adventitial TCs of the perivascular sheaths exhibited strong immunoreactivity to α-SMA. Conversely, despite showing mild immunoreactivity to PDGFRα and CD34, the TCs in normal conjunctiva did not show any immunoreactivity to nestin and VEGF. Moreover, in pterygium and conjunctiva, the TCs were always negative for c-kit., Conclusions: Because of the distribution and immunophenotype, TCs in pterygium may represent a subpopulation of relatively immature cells with regenerative potential. In addition, the expression of nestin may suggest possible involvement of TCs as active players in the regeneration of ultraviolet-damaged stroma and vascular remodeling. The fibrotic transformation in the cicatricial area may stand for a breakdown of the regenerative process.

Pterygium is a common ocular surface disorder characterized by excessive cell proliferation, inflammation, fibrosis, angiogenesis and extracellular matrix remodeling. The Angiotensin converting enzyme (ACE or ACE I) is the major component of the Renin-angiotensin system (RAS) converting the inactive decapeptide Angiotensin I (Ang I) to the active octapeptide Angiotensin II (Ang II). Besides this 'classical role', it can act as transcriptional regulator in response to external stimuli that may lead to cell damage and tissue remodeling. Due to this role, it can be internalized into the nuclear compartment to act as transcriptional factor for proteins involved in the inflammatory response. The aim of the present study was to determine ACE expression and localization in pterygium and culture pterygium cells by immunohistochemistry. Our results are the first to demonstrate nuclear immunolocalization of ACE, more so in pterygium compared to conjunctiva epithelial cells in histological sections. ACE was not detected in the nuclei of subcultivated pterygium epithelial cells. The nuclear localization of ACE may be correlated with an anti-inflammatory path mediated by activation of its transcriptional role.

8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the main mutagenic modifications induced in DNA by oxidative stress. Elevated levels of 8-OHdG have been regarded as an independent prognostic factor in different types of cancer. Various enzymes, such as human 8-oxoguanine DNA-glycosylase 1 (hOGG1) and glucose-6-phosphate dehydrogenase (G6PD), act as protection against oxidative stress. The low activity of such enzymes has been consistently associated with increased risk of progression in several tumor types. The aim of this study was to investigate whether 8-OHdG, hOGG1 and G6PD expression in tumor tissues might be a predictor of survival in melanoma patients. The expression of 8-OHdG, hOGG1 and G6PD was immunohistochemically investigated in primary cutaneous melanoma and the effect on survival was analyzed. Furthermore, the immunostaining for p53 and survivin was evaluated and the relationship among 8-OHdG, hOGG1, G6PD, p53 and survivin expression was analyzed. Kaplan-Meier analysis demonstrated that patients with low expression of nuclear 8-OHdG had significantly longer survival time compared with those with a high expression (P=0.032), whereas cancer-specific survival of patients was not associated with hOGG1 or G6PD expression. These results suggest an involvement of oxidative DNA damage in the process of melanoma pathogenesis and demonstrate that 8-OHdG expression in nuclei of tumor cells could be useful as an early independent prognostic marker in patients with primary cutaneous melanoma.

The cancer stem cell hypothesis suggests that mutated melanocyte stem cells are present in skin as precursors of melanoma cells. Nestin and CD133 have been described as markers of melanocytic stem cells. The aim of this study was to establish if melanocytic stem cells could have a prognostic significance in melanoma progression. An immunohistochemical study for nestin and CD133 was performed in 130 primary tumors and 32 nodal metastasis biopsy specimens to evaluate possible differences, and to compare the results with survival data and clinicopathological variables. Nestin was expressed in cytoplasm of non-pigmented tumor cells and in endothelial cells, especially at the invading tumor front. Nestin staining in stage I and II (according to the American Joint Committee on Cancer Staging system) melanoma patients significantly predicted poor survival (log-rank test, P=0.037), with lower survival rates in cases with nestin positivity in both tumoral and endothelial cells. CD133 staining was not associated with survival. There were no significant differences in nestin or CD133 expression between primary tumors and metastases. These results suggest that nestin expression in both tumoral and endothelial cells may be considered an important early prognostic marker in melanoma.

Purpose: To investigate the expression of cyclooxygenase-2 (COX-2) in a group of 93 Ecuadorian primary pterygia and to evaluate a possible association between COX-2 and survivin., Methods: Primary pterygium samples were treated for the immunohistochemical evaluation of COX-2 and survivin. Mouse monoclonal antibody to COX-2 and rabbit polyclonal antibody to survivin were used. Statistical analysis was performed using the SPSS statistical software package, version 15.0., Results: In our study, 63 (67.7%) primary pterygia samples were positive for COX-2 staining, and 70 (75.3%) specimens were positive for survivin expression. In the group of pterygia with survivin immunostaining, there were 55 (78.6%) samples with COX-2 expression. The staining of both COX-2 and survivin was localized in the lower and middle layers of the epithelium. When analyzed by Fisher's exact test, the expression of COX-2 showed a strong significant correlation with survivin (p=0.0002)., Conclusions: These data, showing a significant correlation between COX-2 and survivin in primary pterygium, suggest that pterygium may originate through an anti-apoptotic mechanism.

The deregulation of apoptosis is characteristic of human carcinogenesis. Survivin, an inhibitor of apoptosis, p53 and p16, two tumour suppressor proteins involved in cell cycle control, play a central role in apoptosis. The aim of this study was to investigate, in primary cutaneous melanoma from 68 patients, the expression of survivin with respect to p53 or p16; the association of these proteins, alone or in combination with clinicopathological features; and, most importantly, to elucidate the role of these markers in predicting survival. The level of survivin expression was significantly higher in the p53 positive group of melanomas compared with the p53 negative one, suggesting a cooperative effect in favouring the progression of melanoma, while no correlation was found between survivin and p16. Moreover, the altered expression of nuclear survivin, p53 and p16 were all associated with poor survival, as demonstrated by univariate analysis. However, these biomarkers have been shown to have superior predictive value when studied in combination (P<0.0001) rather than alone, while the risk of mortality grew progressively with increasing the number of altered biomarkers. These data suggest that the assessment of the combined marker status and number of altered markers in patients with melanoma provides important additional prognostic information that may help in patient selection for adjuvant therapies.