Recent advances on the mechanisms regulating cholangiocyte proliferation and the significance of the neuroendocrine regulation of cholangiocyte pathophysiology

Cholangiocytes are epithelial cells lining the biliary epithelium. Cholangiocytes play several key roles in the modification of ductal bile and are also the target cells in chronic cholestatic liver diseases (i.e., cholangiopathies) such as PSC, PBC, polycystic liver disease (PCLD) and cholangiocarcinoma (CCA). During these pathologies, cholangiocytes (which in normal condition are in a quiescent state) begin to proliferate acquiring phenotypes of neuroendocrine cells, and start secreting different cytokines, growth factors, neuropeptides, and hormones to modulate cholangiocytes proliferation and interaction with the surrounding environment, trying to reestablish the balance between proliferation/loss of cholangiocytes for the maintenance of biliary homeostasis. The purpose of this review is to summarize the recent findings on the mechanisms regulating cholangiocyte proliferation and the significance of the neuroendocrine regulation of cholangiocyte pathophysiology. To clarify the mechanisms of action of these factors we will provide new potential strategies for the management of chronic liver diseases

Microcorrosion Casting in Normal and Pathological Biliary Tree Morphology

The organization of the intrahepatic biliary tree was studied in three dimensions by scanning electron microscopic (SEM) corrosion casts, in normal and cholestatic rat liver. In the normal liver the observation revealed the features of the biliary passages from the bile canaliculi to the canaliculo-ductular junction, to the ductules and the bile ducts, confirming previous SEM observations. In cholestatic liver, the modifications and the proliferation of bile ductules appear clearly. Resin flow from canalicular to sinusoidal network was never observed. The method was found to be very useful in the evaluation of the architecture of the intrahepatic biliary tree, under normal as well as under pathological conditions

Multiple solutions for asteroid orbits: Computational procedure and applications

We describe the Multiple Solutions Method, a one-dimensional sampling of the six-dimensional orbital confidence region that is widely applicable in the field of asteroid orbit determination. In many situations there is one predominant direction of uncertainty in an orbit determination or orbital prediction, i.e., a ``weak'' direction. The idea is to record Multiple Solutions by following this, typically curved, weak direction, or Line Of Variations (LOV). In this paper we describe the method and give new insights into the mathematics behind this tool. We pay particular attention to the problem of how to ensure that the coordinate systems are properly scaled so that the weak direction really reflects the intrinsic direction of greatest uncertainty. We also describe how the multiple solutions can be used even in the absence of a nominal orbit solution, which substantially broadens the realm of applications. There are numerous applications for multiple solutions; we discuss a few problems in asteroid orbit determination and prediction where we have had good success with the method. In particular, we show that multiple solutions can be used effectively for potential impact monitoring, preliminary orbit determination, asteroid identification, and for the recovery of lost asteroids

Common features between neoplastic and preneoplastic lesions of the biliary tract and the pancreas

The bile duct system and pancreas show many similarities due to their anatomical proximity and common embryological origin. Consequently, preneoplastic and neoplastic lesions of the bile duct and pancreas share analogies in terms of molecular, histological and pathophysiological features. Intraepithelial neoplasms are reported in biliary tract, as biliary intraepithelial neoplasm (BilIN), and in pancreas, as pancreatic intraepithelial neoplasm (PanIN). Both can evolve to invasive carcinomas, respectively cholangiocarcinoma (CCA) and pancreatic ductal adenocarcinoma (PDAC). Intraductal papillary neoplasms arise in biliary tract and pancreas. Intraductal papillary neoplasm of the biliary tract (IPNB) share common histologic and phenotypic features such as pancreatobiliary, gastric, intestinal and oncocytic types, and biological behavior with the pancreatic counterpart, the intraductal papillary mucinous neoplasm of the pancreas (IPMN). All these neoplastic lesions exhibit similar immunohistochemical phenotypes, suggesting a common carcinogenic process. Indeed, CCA and PDAC display similar clinic-pathological features as growth pattern, poor response to conventional chemotherapy and radiotherapy and, as a consequence, an unfavorable prognosis. The objective of this review is to discuss similarities and differences between the neoplastic lesions of the pancreas and biliary tract with potential implications on a common origin from similar stem/progenitor cells

H3 histamine receptor-mediated activation of protein kinase calpha inhibits the growth of cholangiocarcinoma in vitro and in vivo

Histamine regulates functions via four receptors (HRH1, HRH2, HRH3, and HRH4). The D-myo-inositol 1,4,5-trisphosphate (IP(3))/Ca(2+)/protein kinase C (PKC)/mitogen-activated protein kinase pathway regulates cholangiocarcinoma growth. We evaluated the role of HRH3 in the regulation of cholangiocarcinoma growth. Expression of HRH3 in intrahepatic and extrahepatic cell lines, normal cholangiocytes, and human tissue arrays was measured. In Mz-ChA-1 cells stimulated with (R)-(alpha)-(-)-methylhistamine dihydrobromide (RAMH), we measured (a) cell growth, (b) IP(3) and cyclic AMP levels, and (c) phosphorylation of PKC and mitogen-activated protein kinase isoforms. Localization of PKC alpha was visualized by immunofluorescence in cell smears and immunoblotting for PKC alpha in cytosol and membrane fractions. Following knockdown of PKC alpha, Mz-ChA-1 cells were stimulated with RAMH before evaluating cell growth and extracellular signal-regulated kinase (ERK)-1/2 phosphorylation. In vivo experiments were done in BALB/c nude mice. Mice were treated with saline or RAMH for 44 days and tumor volume was measured. Tumors were excised and evaluated for proliferation, apoptosis, and expression of PKC alpha, vascular endothelial growth factor (VEGF)-A, VEGF-C, VEGF receptor 2, and VEGF receptor 3. HRH3 expression was found in all cells. RAMH inhibited the growth of cholangiocarcinoma cells. RAMH increased IP(3) levels and PKC alpha phosphorylation and decreased ERK1/2 phosphorylation. RAMH induced a shift in the localization of PKC alpha expression from the cytosolic domain into the membrane region of Mz-ChA-1 cells. Silencing of PKC alpha prevented RAMH inhibition of Mz-ChA-1 cell growth and ablated RAMH effects on ERK1/2 phosphorylation. In vivo, RAMH decreased tumor growth and expression of VEGF and its receptors; PKC alpha expression was increased. RAMH inhibits cholangiocarcinoma growth by PKC alpha-dependent ERK1/2 dephosphorylation. Modulation of PKC alpha by histamine receptors may be important in regulating cholangiocarcinoma growth. (Mol Cancer Res 2009;7(10):1704-13

Endoscopic Pilonidal Sinus Treatment. A Tertiary Care Academic Center Experience

Background: Pilonidal disease (PD) represents one of the most common proctological diseases in young adults. Although several approaches to treating PD have been described, there is still a lack of agreement on which is the best. The aim of this study was to evaluate the long-term efficacy of endoscopic pilonidal sinus treatment (EPSiT) at a tertiary care academic center. Methods: Between June 2017 and January 2021, a total of 32 patients [12 women (37.5%) and 20 men (62.5%)] with a mean age of 29.22 ± 12.98 years were treated with EPSiT. Pre- and post-operative symptoms were assessed with a score of 0–5. Success was defined as the absence of any subjective symptoms, as well as by complete post-operative wound healing. Results: Most of the patients had a midline external opening (17/32; 53.1%), with a mean number of external openings of 2.41 (1–4) ± 1.04. The median post-operative pain score was 0, and the mean follow-up period was 22 (4–42) ± 11.49 months. The time to wound healing was reduced in patients with one opening (28.14 ± 4.06 days) compared to patients with two or more openings (33.64 ± 7.3 days) (p = 0.067). The mean operative time was longer in patients who subsequently had a recurrence (41.75 ± 6.24 vs. 34.18 ± 6.24 min; p = 0.031). The overall success rate was 87.5% (28/32), and the mean time to recurrence was 3.25 (2–5) ± 1.26 months. Conclusions: EPSiT represents a viable option for the treatment of PD. More evidence and a longer follow-up period are needed to validate the results

Cell Therapy and Bioengineering in Experimental Liver Regenerative Medicine: In Vivo Injury Models and Grafting Strategies

Abstract Purpose of Review To describe experimental liver injury models used in regenerative medicine, cell therapy strategies to repopulate damaged livers and the efficacy of liver bioengineering. Recent Findings Several animal models have been developed to study different liver conditions. Multiple strategies and modified protocols of cell delivery have been also reported. Furthermore, using bioengineered liver scaffolds has shown promising results that could help in generating a highly functional cell delivery system and/or a whole transplantable liver. Summary To optimize the most effective strategies for liver cell therapy, further studies are required to compare among the performed strategies in the literature and/or innovate a novel modifying technique to overcome the potential limitations. Coating of cells with polymers, decellularized scaffolds, or microbeads could be the most appropriate solution to improve cellular efficacy. Besides, overcoming the problems of liver bioengineering may offer a radical treatment for end-stage liver diseases

Anatomical, histomorphological and molecular classification of cholangiocarcinoma

Cholangiocarcinoma constitutes a heterogeneous group of malignancies that can emerge at any point of the biliary tree. Cholangiocarcinoma is classified into intrahepatic, perihilar and distal based on its anatomical location. Histologically, conventional perihilar/distal cholangiocarcinomas are mucin-producing adenocarcinomas or papillary tumours; intrahepatic cholangiocarcinomas are more heterogeneous and can be sub-classified according to the level or size of the displayed bile duct. Cholangiocarcinoma develops through multistep carcinogenesis and is preceded by dysplastic and in situ lesions. Definition and clinical significance of precursor lesions, including biliary intraepithelial neoplasia, intraductal papillary neoplasms of the bile duct, intraductal tubulopapillary neoplasms and mucinous cystic neoplasm, are discussed in this review. A main challenge in diagnosing cholangiocarcinoma is the fact that tumour tissue for histological examination is difficult to obtain. Thus, a major clinical obstacle is the establishment of the correct diagnosis at a tumour stage that is amenable to surgery which still represents the only curable therapeutic option. Current standards, methodology and criteria for diagnosis are discussed. Cholangiocarcinoma represents a heterogeneous tumour with regard to molecular alterations. In intrahepatic subtype, mainly two distinctive morpho-molecular groups can currently be discriminated. Large-duct type intrahepatic cholangiocarcinoma shows a high mutation frequency of oncogenes and tumour suppressor genes, such as KRAS and TP53 while Isocitrate Dehydrogenase 1/2 mutations and Fibroblast Growth Factor Receptor 2-fusions are typically seen in small-duct type tumours. It is most important to ensure the separation of the given anatomical subtypes and to search for distinct subgroups within the subtypes on a molecular and morphological basis

Composition of Ragusano Cheese During Aging

Ragusano cheese is a brine-salted pasta filata cheese. Composition changes during 12 mo of aging were determined. Historically, Ragusano cheese has been aged in caves at 14 to 16 degrees C with about 80 to 90% relative humidity. Cheeses (n = 132) included in our study of block-to-block variation were produced by 20 farmhouse cheese makers in the Hyblean plain region of the Province of Ragusa in Sicily. Mean initial cheese block weight was about 14 kg. The freshly formed blocks of cheese before brine salting contained about 45.35% moisture, 25.3% protein, and 25.4% fat, with a pH of 5.25. As result of the brining and aging process, a natural rind forms. After 12 mo of aging, the cheese contained about 33.6% moisture, 29.2% protein, 30.0% fat, and 4.4% salt with a pH of 5.54, but block-to-block variation was large. Both soluble nitrogen content and free fatty acid (FFA) content increased with age. The pH 4.6 acetate buffer and 12% TCA-soluble nitrogen as a percentage of total nitrogen were 16 and 10.7%, respectively, whereas the FFA content was about 643 mg/100 g of cheese at 180 d. Five blocks of cheese were selected at 180 d for a study of variation within block. Composition variation within block was large; the center had higher moisture and lower salt in moisture content than did the outside. Composition variation within blocks favored more proteolysis and softer texture in the center

The FXR agonist obeticholic acid inhibits the cancerogenic potential of human cholangiocarcinoma

Cholangiocarcinoma (CCA) is an aggressive cancer with high resistance to chemotherapeutics. CCA is enriched in cancer stem cells, which correlate with aggressiveness and prognosis. FXR, a member of the metabolic nuclear receptor family, is markedly down-regulated in human CCA. Our aim was to evaluate, in primary cultures of human intrahepatic CCA (iCCA), the effects of the FXR agonist obeticholic acid (OCA), a semisynthetic bile acid derivative, on their cancerogenic potential. Primary human iCCA cell cultures were prepared from surgical specimens of mucinous or mixed iCCA subtypes. Increasing concentrations (0–2.5 μM) of OCA were added to culture media and, after 3–10 days, effects on proliferation (MTS assay, cell population doubling time), apoptosis (annexin V-FITC/propidium iodide), cell migration and invasion (wound healing response and Matrigel invasion assay), and cancerogenic potential (spheroid formation, clonogenic assay, colony formation capacity) were evaluated. Results: FXR gene expression was downregulated (RT-qPCR) in iCCA cells vs normal human biliary tree stem cells (p < 0.05) and in mucinous iCCA vs mixed iCCA cells (p < 0.05) but was upregulated by addition of OCA. OCA significantly (p < 0.05) inhibited proliferation of both mucinous and mixed iCCA cells, starting at a concentration as low as 0.05 μM. Also, CDCA (but not UDCA) inhibited cell proliferation, although to a much lower extent than OCA, consistent with its different affinity for FXR. OCA significantly induced apoptosis of both iCCA subtypes and decreased their in vitro cancerogenic potential, as evaluated by impairment of colony and spheroid formation capacity and delayed wound healing and Matrigel invasion. In general, these effects were more evident in mixed than mucinous iCCA cells. When tested together with Gemcitabine and Cisplatin, OCA potentiated the anti-proliferative and pro-apoptotic effects of these chemotherapeutics, but mainly in mixed iCCA cells. OCA abolished the capacity of both mucinous and mixed iCCA cells to form colonies when administered together with Gemcitabine and Cisplatin. In subcutaneous xenografts of mixed iCCA cells, OCA alone or combined with Gemcitabine or Cisplatin markedly reduced the tumor size after 5 weeks of treatment by inducing necrosis of tumor mass and inhibiting cell proliferation. In conclusion, FXR is down-regulated in iCCA cells, and its activation by OCA results in anti-cancerogenic effects against mucinous and mixed iCCA cells, both in vitro and in vivo. The effects of OCA predominated in mixed iCCA cells, consistent with the lower aggressiveness and the higher FXR expression in this CCA subtype. These results, showing the FXR-mediated capacity of OCA to inhibit cholangiocarcinogenesis, represent the basis for testing OCA in clinical trials of CCA patients