• Article hsp60 quantification in human gastric mucosa shows differences between pathologies with various degrees of proliferation and malignancy grade

      Pitruzzella, Alessandro; Burgio, Stefano; Lo Presti, Pietro; Ingrao, Sabrina; Fucarino, Alberto; Bucchieri, Fabio; Cabibi, Daniela; Cappello, Francesco; Conway de Macario, Everly; Macario, Alberto J.L.; et al. (MDPI AG, 2021-04-16)
      Background: Stomach diseases are an important sector of gastroenterology, including proliferative benign; premalignant; and malignant pathologies of the gastric mucosa, such as gastritis, hyperplastic polyps, metaplasia, dysplasia, and adenocarcinoma. There are data showing quantitative changes in chaperone system (CS) components in inflammatory pathologies and tumorigenesis, but their roles are poorly understood, and information pertaining to the stomach is scarce. Here, we report our findings on one CS component, the chaperone Hsp60, which we studied first considering its essential functions inside and outside mitochondria. Methods: We performed immunohistochemical experiments for Hsp60 in different samples of gastric mucosa. Results: The data obtained by quantitative analysis showed that the average percentages of Hsp60 were of 32.8 in normal mucosa; 33.5 in mild-to-moderate gastritis; 51.8 in severe gastritis; 58.5 in hyperplastic polyps; 67.0 in intestinal metaplasia; 89.4 in gastric dysplasia; and 92.5 in adenocarcinomas. Noteworthy were: (i) the difference between dysplasia and adenocarcinoma with the other pathologies; (ii) the progressive increase in Hsp60 from gastritis to hyperplastic polyp, gastric dysplasia, and gastric carcinoma; and (iii) the correlation of Hsp60 levels with histological patterns of cell proliferation and, especially, with tissue malignancy grades. Conclusions: This trend likely reflects the mounting need for cells for Hsp60 as they progress toward malignancy and is a useful indicator in differential diagnosis, as well as the call for research on the mechanisms underpinning the increase in Hsp60 and its possible roles in carcinogenesis. © 2021 by the authors.
    • The chaperone system in glioblastoma multiforme and derived cell lines: diagnostic and mechanistic implications.

      Alberti, Giusi; Campanella, Claudia; Paladino, Letizia; Porcasi, Rossana; Bavisotto, Celeste Caruso; Pitruzzella, Alessandro; Graziano, Francesca; Florena, Ada Maria; Argo, Antonina; de Macario, Everly Conway; et al. (IMR Press, 2022-03-15)
      BACKGROUND: Glioblastoma multiforme (GBM) is the most common and malignant primary brain tumor in adults. Novel treatments are needed to counteract the molecular mechanisms of GBM growth and drug resistance. The chaperone system (CS) members are typically cytoprotective but some, termed Hsp, can become pathogenic and participate in carcinogenesis, along with the vascular endothelial growth factor (VEGF), and we investigated them in GBM biopsies and derived cell lines. The objectives were to identify diagnostic-prognostic biomarkers and gather information for developing chaperonotherapy. METHODS: Cell lines from GBMs were established, characterized (morphology, growth characteristics, and specific markers), and stored. Chaperones and angiogenic factors [Hsp10, Hsp27, Hsp60, Hsp70, Hsp90, FLT-1 (VEGFR-1), FLK1 (KDR, VEGFR-2), and FLT-4 (VEGFR-3)] were observed in cells by immunofluorescence while the chaperones were measured in tumor tissue by immunohistochemistry. RESULTS: Four cell lines were derived from four different GBMs; the cells were spindle shaped or polygonal and grew at high rates as adherent monolayers or clusters without evidence of contact inhibition. The astrocyte-specific glial fibrillary acidic protein (GFAP); and the neuronal NSE, malignancy VIM, and proliferation PCNA, markers were determined. The cells expressed GFAP but no NSE, indicating that they were primary glioblastoma cell lines, with high levels of Hsp10, Hsp27, Hsp60, Hsp90, and Flk1; and low levels of Hsp70, Flt1, and Flt4. CONCLUSIONS: Four cell lines were established derived from four out of ten GBM tumors studied. The cell lines showed intense positivity for chaperones studied and factors connected to malignancy and the tumors showed increased levels of chaperones, making them potential diagnostic-prognostic biomarkers and targets for anti-cancer compounds.
    • Molecular Chaperones and Thyroid Cancer

      Paladino, Letizia; Vitale, Alessandra Maria; Santonocito, Radha; Pitruzzella, Alessandro; Cipolla, Calogero; Graceffa, Giuseppa; Bucchieri, Fabio; Conway de Macario, Everly; Macario, Alberto J L; Rappa, Francesca (MDPI AG, 2021-04-18)
      Thyroid cancers are the most common of the endocrine system malignancies and progress must be made in the areas of differential diagnosis and treatment to improve patient management. Advances in the understanding of carcinogenic mechanisms have occurred in various fronts, including studies of the chaperone system (CS). Components of the CS are found to be quantitatively increased or decreased, and some correlations have been established between the quantitative changes and tumor type, prognosis, and response to treatment. These correlations provide the basis for identifying distinctive patterns useful in differential diagnosis and for planning experiments aiming at elucidating the role of the CS in tumorigenesis. Here, we discuss studies of the CS components in various thyroid cancers (TC). The chaperones belonging to the families of the small heat-shock proteins Hsp70 and Hsp90 and the chaperonin of Group I, Hsp60, have been quantified mostly by immunohistochemistry and Western blot in tumor and normal control tissues and in extracellular vesicles. Distinctive differences were revealed between the various thyroid tumor types. The most frequent finding was an increase in the chaperones, which can be attributed to the augmented need for chaperones the tumor cells have because of their accelerated metabolism, growth, and division rate. Thus, chaperones help the tumor cell rather than protect the patient, exemplifying chaperonopathies by mistake or collaborationism. This highlights the need for research on chaperonotherapy, namely the development of means to eliminate/inhibit pathogenic chaperones.
    • Muscle Histopathological Abnormalities in a Patient With a CCT5 Mutation Predicted to Affect the Apical Domain of the Chaperonin Subunit.

      Scalia, Federica; Barone, Rosario; Rappa, Francesca; Marino Gammazza, Antonella; Lo Celso, Fabrizio; Lo Bosco, Giosuè; Barone, Giampaolo; Antona, Vincenzo; Vadalà, Maria; Vitale, Alessandra Maria; et al. (Frontiers Media S.A., 2022-06-02)
      Recognition of diseases associated with mutations of the chaperone system genes, e.g., chaperonopathies, is on the rise. Hereditary and clinical aspects are established, but the impact of the mutation on the chaperone molecule and the mechanisms underpinning the tissue abnormalities are not. Here, histological features of skeletal muscle from a patient with a severe, early onset, distal motor neuropathy, carrying a mutation on the CCT5 subunit (MUT) were examined in comparison with normal muscle (CTR). The MUT muscle was considerably modified; atrophy of fibers and disruption of the tissue architecture were prominent, with many fibers in apoptosis. CCT5 was diversely present in the sarcolemma, cytoplasm, and nuclei in MUT and in CTR and was also in the extracellular space; it colocalized with CCT1. In MUT, the signal of myosin appeared slightly increased, and actin slightly decreased as compared with CTR. Desmin was considerably delocalized in MUT, appearing with abnormal patterns and in precipitates. Alpha-B-crystallin and Hsp90 occurred at lower signals in MUT than in CTR muscle, appearing also in precipitates with desmin. The abnormal features in MUT may be the consequence of inactivity, malnutrition, denervation, and failure of protein homeostasis. The latter could be at least in part caused by malfunction of the CCT complex with the mutant CCT5 subunit. This is suggested by the results of the in silico analyses of the mutant CCT5 molecule, which revealed various abnormalities when compared with the wild-type counterpart, mostly affecting the apical domain and potentially impairing chaperoning functions. Thus, analysis of mutated CCT5 in vitro and in vivo is anticipated to provide additional insights on subunit involvement in neuromuscular disorders.