Total and caspase-cleaved cytokeratin 18 in chronic cholecystitis: A prospective study

<p>Abstract</p> <p>Background</p> <p>Cell death mode has been studied in cancer, autoimmune, and neurodegenerative diseases. In this study, apoptosis and necrosis are investigated for the first time in patients with chronic calculous cholecystitis.</p> <p>Methods and materials</p> <p>Thirty five (35) patients (27 women and 8 men, aged 55.65 ± 13.48 years) with symptomatic chronic calculous cholecystitis underwent laparoscopic cholecystectomy. The early specific apoptotic tendency (caspase-cleaved cytokeratin 18) was studied in these patients with M30 Apoptosense ELISA and the total cytokerarin 18 (both derived from apoptosis and necrosis) with M65 ELISA. The ratio M30/M65 (caspase-cleaved to total cytokeratin 18) was also computed. According to the histopathological examination, the patients were divided in two groups: group A included patients with chronic inactive cholecystitis (n = 10), and group B those with chronic active cholecystitis (n = 25).</p> <p>Results</p> <p>The concentrations of caspase-cleaved cytokerarin 18 (CK18), and especially those of total CK18, were higher in bile samples than in serum samples. In group B, there were significant differences between serum and bile samples regarding both caspase-cleaved CK18 and total CK18. Cells staining positive for caspase-cleaved CK18 were present in the epithelial cells of the mucosa of the gallbladder.</p> <p>Conclusion</p> <p>CK18 is expressed in the gallbladder epithelial cells. The concentrations of both caspase-cleaved CK18 and total CK18 were higher in bile samples than in serum samples. The levels of total CK18, as well as caspase-cleaved CK18, do not seem to differ between active and inactive chronic cholecystitis.</p

Primary Malignant Melanoma of the Cervix: A Case Report and a Review of the Literature

Background. Gynecologic melanomas are extremely rare malignancies, and primary malignant melanoma of the cervix (PMMC) is the rarest among them all, with less than 100 cases reported so far. Although some conditions have been correlated with the pathogenesis of this entity, no specific risk factor has been yet identified, with vaginal bleeding being the most common symptoms. The diagnosis is based on physical examination with speculum assessment and cytologic and histopathologic findings accompanied with immunohistochemical staining of lesion’s biopsies. Case Presentation. We report a case of PMMC in a 34-year-old para-2 patient, among the youngest cases of PMMC reported, that presented to our clinic for routine examination. Gynecologic examination demonstrated a dark, heavily fully pigmented cervical growth completely covering the entire external cervical os. Biopsy obtained and showed malignant melanoma. She underwent radical hysterectomy with bilateral salpingo-oophorectomy and pelvic lymphadenectomy. The pathological diagnosis was FIGO stage IB1 PMMC. Despite 2 courses of anti-PD-1 antibody (Nivolumab) treatment, the patient passed away 13 months after diagnosis (12 months after surgery). Conclusions. Early diagnosis and subsequently early treatment are of high importance regarding patients’ prognosis and survival. No standardized protocols or treatment guidelines specific for this rare cancer have been issued; thus, clinicians are called to assess each case individually. Current treatment options are based on surgical excision mostly with radical hysterectomy, but in advanced or recurrent state of the disease, other treatment modalities, such as chemotherapy, radiotherapy, and immunotherapy, can be employed. Prognosis for these patients is very poor, and survival rate remains extremely low, with the median OS reported being less than 2 years. Reporting and publishing of such cases are both of paramount importance for the better understanding of this uncommon cervical malignancy, and further biological and clinical investigations are required for more suitable and effective therapies to be determined. A new staging system, specific to PMMC, could be of great use for the better correlation of the disease’s stage and prognosis of these patients

The Effect of Strontium Ranelate on Fracture Healing: An Animal Study

Background. Strontium ranelate (StR) is an antiosteoporotic agent previously utilized for the enhancement of fracture union. We investigated the effects of StR on fracture healing using a rabbit model. Methods. Forty adult female rabbits were included in the study and were divided in 2 equal groups, according to StR treatment or untreated controls. All animals were subjected to osteotomy of the ulna, while the contralateral ulna remained intact and served as a control for the biomechanical assessment of fracture healing. Animals in the study group received 600 mg/kg/day of StR orally. All animals received ordinary food. At 2 and 4 weeks, all animals were euthanatized and the osteotomy sites were evaluated for healing through radiological, biomechanical, and histopathological studies. Results. The treatment group presented statistically significant higher callus diameter, total callus area, percentage of fibrous tissue (p<0.001), vessels/mm2, number of total vessels, and lower osteoclast number/mm2 (p<0.05) than the control group at 2 weeks. Additionally, the treatment group presented significantly higher percentages of new trabecular bone, vessels/mm2, osteoclast number/mm2, and lower values for callus diameter, as well as total callus area (p<0.05), than the control group at 4 weeks. At 4 weeks, in the treatment group, force applied (p=0.003), energy at failure (p=0.004), and load at failure (p=0.003) were all significantly higher in the forearm specimens with the osteotomized ulnae compared to those without. Radiological bone union was demonstrated for animals receiving StR at 4 weeks compared with controls (p=0.045). Conclusion. StR appears to enhance fracture healing but further studies are warranted in order to better elucidate the mechanisms and benefits of StR treatment

The Effect of Strontium Ranelate on Fracture Healing: An Animal Study

Background. Strontium ranelate (StR) is an antiosteoporotic agent previously utilized for the enhancement of fracture union. We investigated the effects of StR on fracture healing using a rabbit model. Methods. Forty adult female rabbits were included in the study and were divided in 2 equal groups, according to StR treatment or untreated controls. All animals were subjected to osteotomy of the ulna, while the contralateral ulna remained intact and served as a control for the biomechanical assessment of fracture healing. Animals in the study group received 600 mg/kg/day of StR orally. All animals received ordinary food. At 2 and 4 weeks, all animals were euthanatized and the osteotomy sites were evaluated for healing through radiological, biomechanical, and histopathological studies. Results. The treatment group presented statistically significant higher callus diameter, total callus area, percentage of fibrous tissue (p&lt;0.001), vessels/mm(2), number of total vessels, and lower osteoclast number/mm(2) (p&lt;0.05) than the control group at 2 weeks. Additionally, the treatment group presented significantly higher percentages of new trabecular bone, vessels/mm(2), osteoclast number/mm(2), and lower values for callus diameter, as well as total callus area (p&lt;0.05), than the control group at 4 weeks. At 4 weeks, in the treatment group, force applied (p=0.003), energy at failure (p=0.004), and load at failure (p=0.003) were all significantly higher in the forearm specimens with the osteotomized ulnae compared to those without. Radiological bone union was demonstrated for animals receiving StR at 4 weeks compared with controls (p=0.045). Conclusion. StR appears to enhance fracture healing but further studies are warranted in order to better elucidate the mechanisms and benefits of StR treatment

Transcriptional Regulation of Serine/Threonine Kinase-15 (STK15) Expression by Hypoxia and HIF-1

The serine/threonine kinase-15 (STK15) acts as a cell cycle regulator being overexpressed in various tumors. One mechanism that could contribute to overexpression of STK15 is tumor hypoxia where hypoxia-inducible factor-1 (HIF-1) is a major regulator of transcription. Therefore, we analyzed whether hypoxia and HIF-1 could contribute to overexpression of STK15. We found that hypoxia increased STK15 expression and STK15 promoter activity in HepG2 tumor cells. Overexpression of HIF-1α induced STK15 gene transcription, whereas HIF-1α siRNA and overexpression of prolyl hydroxylase 2 (PHD-2), a negative regulator of HIF-1α, reversed this effect. In addition, site-directed mutagenesis experiments and chromatin immunoprecipitation revealed that from the three putative hypoxia responsive elements (HRE) within the STK15 promoter only HRE-2 was functional and bound HIF-1. Further, siRNA against STK15 inhibited proliferation of HepG2 cells induced by hypoxia. These results show that STK15 gene transcription can be regulated by hypoxia and HIF-1 via HRE-2 of the STK15 promoter. Thus, tumor hypoxia may trigger overexpression of STK15 observed in various tumors

Guidelines for the use and interpretation of assays for monitoring autophagy

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field