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Background: Chronic lateral epicondylitis (LE), also known as tennis elbow, affects 1–3% of the population, primarily those over 40 years old. Most cases resolve with conservative treatments, but some require more advanced interventions. Extracorporeal shockwave therapy (ESWT) has emerged as a non-surgical treatment option, utilizing either low- or high-energy levels to alleviate pain and improve function. Objective: This study aimed to compare the efficacy of low-energy versus high-energy ESWT in the treatment of chronic LE, focusing on pain relief and functional improvement. Methods: A retrospective observational study was conducted including patients treated for chronic LE between 2021 and 2024. Participants were divided into two groups: low-energy ESWT (0.10 mJ/mm2) and high-energy ESWT (0.20 mJ/mm2). Both groups received 2400 pulses at a frequency of 6 Hz once a week for three weeks. Pain and functional outcomes were measured using a visual analog scale (VAS) and the Patient-Rated Tennis Elbow Evaluation Questionnaire (PRTEE) at the baseline, three months (T1), and six months (T2) post-treatment. Results: Forty-six patients participated, with 24 in the low-energy group and 22 in the high-energy group. Baseline demographics and clinical characteristics were similar across groups. At T1 and T2, the low-energy group showed significantly greater reductions in the VAS scores (T1: 4.45 ± 0.8 vs. 3.6 ± 1.7, p = 0.04; T2: 3.2 ± 1.2 vs. 2.1 ± 1.1, p = 0.004) and PRTEE scores (T1: 34.3 ± 6.9 vs. 26.8 ± 11.9, p = 0.03; T2: 25.3 ± 6 vs. 17.6 ± 9, p = 0.005). Significant treatment–time interactions were observed for both the VAS and PRTEE scores, indicating sustained improvements in the low-energy group. Conclusions: Low-energy ESWT was more effective than high-energy ESWT in treating chronic LE, providing greater and longer-lasting pain relief and functional improvement. These findings suggest that low-energy ESWT should be preferred in clinical practice for managing this condition. Future research should focus on larger sample sizes and randomized controlled trials to confirm these results and explore the underlying mechanisms of differential efficacy between energy levels.

Cholangiocytes, the epithelial cells that line the biliary tree, can proliferate under the stimulation of several factors through both autocrine and paracrine pathways. The cocaine-amphetamine-regulated-transcript (CART) peptide has several physiological functions, and it is widely expressed in several organs. CART increases the survival of hippocampal neurons by upregulating brain-derived neurotrophic factor (BDNF), whose expression has been correlated to the proliferation rate of cholangiocytes. In the present study, we aimed to evaluate the expression of CART and its role in modulating cholangiocyte proliferation in healthy and bile duct ligated (BDL) rats in vivo, as well as in cultured normal rat cholangiocytes (NRC) in vitro. Liver samples from both healthy and BDL (1 week) rats, were analyzed by immunohistochemistry and immunofluorescence for CART, CK19, TrkB and p75NTR BDNF receptors. PCNA staining was used to evaluate the proliferation of the cholangiocytes, whereas TUNEL assay was used to evaluate biliary apoptosis. NRC treated or not with CART were used to confirm the role of CART on cholangiocytes proliferation and the secretion of BDNF. Cholangiocytes proliferation, apoptosis, CART and TrkB expression were increased in BDL rats, compared to control rats. We found a higher expression of TrkB and p75NTR, which could be correlated with the proliferation rate of biliary tree during BDL. The in vitro study demonstrated increased BDNF secretion by NRC after treatment with CART compared with control cells. As previously reported, proliferating cholangiocytes acquire a neuroendocrine phenotype, modulated by several factors, including neurotrophins. Accordingly, CART may play a key role in the remodeling of biliary epithelium during cholestasis by modulating the secretion of BDNF.

Abstract Background Alcohol-related liver disease (ALD) is characterized by ductular reaction (DR), liver inflammation, steatosis, fibrosis, and cirrhosis. The secretin (Sct)/secretin receptor (SR) axis (expressed only by cholangiocytes) regulates liver phenotypes in cholestasis. We evaluated the role of Sct signaling on ALD phenotypes. Methods We used male wild-type and Sct−/− mice fed a control diet (CD) or ethanol (EtOH) for 8 wk. Changes in liver phenotypes were measured in mice, female/male healthy controls, and patients with alcoholic cirrhosis. Since Cyp4a10 and Cyp4a11/22 regulate EtOH liver metabolism, we measured their expression in mouse/human liver. We evaluated: (i) the immunoreactivity of the lipogenesis enzyme elongation of very-long-chain fatty acids 1 (Elovl, mainly expressed by hepatocytes) in mouse/human liver sections by immunostaining; (ii) the expression of miR-125b (that is downregulated in cholestasis by Sct) in mouse liver by qPCR; and (iii) total bile acid (BA) levels in mouse liver by enzymatic assay, and the mRNA expression of genes regulating BA synthesis (cholesterol 7a-hydroxylase, Cyp27a1, 12a-hydroxylase, Cyp8b1, and oxysterol 7a-hydroxylase, Cyp7b11) and transport (bile salt export pump, Bsep, Na+-taurocholate cotransporting polypeptide, NTCP, and the organic solute transporter alpha (OSTa) in mouse liver by qPCR. Results In EtOH-fed WT mice there was increased biliary and liver damage compared to control mice, but decreased miR-125b expression, phenotypes that were blunted in EtOH-fed Sct−/− mice. The expression of Cyp4a10 increased in cholangiocytes and hepatocytes from EtOH-fed WT compared to control mice but decreased in EtOH-fed Sct−/− mice. There was increased immunoreactivity of Cyp4a11/22 in patients with alcoholic cirrhosis compared to controls. The expression of miR-125b decreased in EtOH-fed WT mice but returned at normal values in EtOH-fed Sct−/− mice. Elovl1 immunoreactivity increased in patients with alcoholic cirrhosis compared to controls. There was no difference in BA levels between WT mice fed CD or EtOH; BA levels decreased in EtOH-fed Sct−/− compared to EtOH-fed WT mice. There was increased expression of Cyp27a1, Cyp8b1, Cyp7b1, Bsep, NTCP and Osta in total liver from EtOH-fed WT compared to control mice, which decreased in EtOH-fed Sct−/− compared to EtOH-fed WT mice. Conclusions Targeting Sct/SR signaling may be important for modulating ALD phenotypes.

Contrasting evidence is present regarding the contribution of stem/progenitor cell populations to pancreatic regeneration in diabetes. Interestingly, a cell compartment with stem/progenitor cell features has been identified in the pancreatic duct glands (PDGs). The aims of the present study were to evaluate pancreatic islet injury and regeneration, and the participation of the PDG compartment in type 2 diabetic mellitus (T2DM) and in an experimental model of diabetes. Human pancreata were obtained from normal (N = 5) or T2DM (N = 10) cadaveric organ donors. Experimental diabetes was generated in mice by intraperitoneal injection of 150 mg/kg of streptozotocin (STZ, N = 10); N = 10 STZ mice also received daily intraperitoneal injections of 100 µg of human recombinant PDX1 peptide (STZ + PDX1). Samples were examined by immunohistochemistry/immunofluorescence or RT-qPCR. Serum glucose and c-peptide levels were measured in mice. Islets in T2DM patients showed β-cell loss, signs of injury and proliferation, and a higher proportion of central islets. PDGs in T2DM patients had a higher percentage of proliferating and insulin+ or glucagon+ cells compared to controls; pancreatic islets could be observed within pancreatic duct walls of T2DM patients. STZ mice were characterized by reduced islet area compared to controls. PDX1 treatment increased islet area and the percentage of central islets compared to untreated STZ mice but did not revert diabetes. In conclusion, T2DM patients show signs of pancreatic islet regeneration and involvement of the PDG niche. PDX1 administration could support increased endocrine pancreatic regeneration in STZ. These findings contribute to defining the role and participation of stem/progenitor cell compartments within the pancreas.

Liver cancer represents a global health challenge with worldwide growth. Hepatocellular carcinoma (HCC) is the most common type of liver cancer. Indeed, approximately 90% of HCC cases have a low survival rate. Moreover, cholangiocarcinoma (CC) is another malignant solid tumor originating from cholangiocytes, the epithelial cells of the biliary system. It is the second-most common primary liver tumor, with an increasing course in morbidity and mortality. Tumor cells always show high metabolic levels, antioxidant modifications, and an increased iron uptake to maintain unlimited growth. In recent years, alterations in iron metabolism have been shown to play an important role in the pathogenesis of HCC. Several findings show that a diet rich in iron can enhance HCC risk. Hence, elevated iron concentration inside the cell may promote the development of HCC. Growing evidence sustains that activating ferroptosis may potentially block the proliferation of HCC cells. Even in CC, it has been shown that ferroptosis plays a crucial role in the treatment of tumors. Several data confirmed the inhibitory effect in cell growth of photodynamic therapy (PDT) that can induce reactive oxygen species (ROS) in CC, leading to an increase in malondialdehyde (MDA) and a decrease in intracellular glutathione (GSH). MDA and GSH depletion/modulation are crucial in inducing ferroptosis, suggesting that PDT may have the potential to induce this kind of cell death through these ways. A selective induction of programmed cell death in cancer cells is one of the main treatments for malignant tumors; thus, ferroptosis may represent a novel therapeutic strategy against HCC and CC.

Background & aims: Cholangiocytes are the target cells of liver diseases that are characterized by biliary senescence (evidenced by enhanced levels of senescence-associated secretory phenotype, SASP, e.g., TGF-β1), and liver inflammation and fibrosis accompanied by altered bile acid (BA) homeostasis. Taurocholic acid (TC) stimulates biliary hyperplasia by activation of 3′,5′-cyclic cyclic adenosine monophosphate (cAMP) signaling, thereby preventing biliary damage (caused by cholinergic/adrenergic denervation) through enhanced liver angiogenesis. Also: (i) α-calcitonin gene-related peptide (α-CGRP, which activates the calcitonin receptor-like receptor, CRLR), stimulates biliary proliferation/senescence and liver fibrosis by enhanced biliary secretion of SASPs; and (ii) knock-out of α-CGRP reduces these phenotypes by decreased cAMP levels in cholestatic models. We aimed to demonstrate that TC effects on liver phenotypes are dependent on changes in the α-CGRP/CALCRL/cAMP/PKA/ERK1/2/TGF-β1/VEGF axis. Methods: Wild-type and α-CGRP−/− mice were fed with a control (BAC) or TC diet for 1 or 2 wk. We measured: (i) CGRP levels by both ELISA kits in serum and by qPCR in isolated cholangiocytes (CALCA gene for α-CGRP); (ii) CALCRL immunoreactivity by immunohistochemistry (IHC) in liver sections; (iii) liver histology, intrahepatic biliary mass, biliary senescence (by β-GAL staining and double immunofluorescence (IF) for p16/CK19), and liver fibrosis (by Red Sirius staining and double IF for collagen/CK19 in liver sections), as well as by qPCR for senescence markers in isolated cholangiocytes; and (iv) phosphorylation of PKA/ERK1/2, immunoreactivity of TGF-β1/TGF- βRI and angiogenic factors by IHC/immunofluorescence in liver sections and qPCR in isolated cholangiocytes. We measured changes in BA composition in total liver by liquid chromatography/mass spectrometry. Results: TC feeding increased CALCA expression, biliary damage, and liver inflammation and fibrosis, as well as phenotypes that were associated with enhanced immunoreactivity of the PKA/ERK1/2/TGF-β1/TGF-βRI/VEGF axis compared to BAC-fed mice and phenotypes that were reversed in α-CGRP−/− mice fed TC coupled with changes in hepatic BA composition. Conclusion: Modulation of the TC/ α-CGRP/CALCRL/PKA/ERK1/2/TGF-β1/VEGF axis may be important in the management of cholangiopathies characterized by BA accumulation.

Cholangiocarcinoma (CCA) represents the second most common primary hepatic malignancy and originates from the neoplastic transformation of the biliary cells. The intrahepatic subtype includes two morpho-molecular forms: large-duct type intrahepatic CCA (iCCA) and small-duct type iCCA. Iron is fundamental for the cellular processes, contributing in tumor development and progression. The aim of this study was to evaluate iron uptake, storage, and efflux proteins in both lipopolysaccharide-inflamed small and large cholangiocytes as well as in different iCCA subtypes. Our results show that, despite an increase in interleukin-6 production by both small and large cholangiocytes, ferroportin (Fpn) was decreased only in small cholangiocytes, whereas transferrin receptor-1 (TfR1) and ferritin (Ftn) did not show any change. Differently from in vitro models, Fpn expression was increased in malignant cholangiocytes of small-duct type iCCA in comparison to large-duct type iCCA and peritumoral tissues. TfR1, Ftn and hepcidin were enhanced, even if at different extent, in both malignant cholangiocytes in comparison to the surrounding samples. Lactoferrin was higher in large-duct type iCCA in respect to small-duct type iCCA and peritumoral tissues. These findings show a different iron handling by inflamed small and large cholangiocytes, and small and large-duct type iCCA. The difference in iron homeostasis by the iCCA subtypes may have implications for the tumor management.

In normal human livers, EpCAMpos cells are mostly restricted in two distinct niches, which are (i) the bile ductules and (ii) the mucous glands present inside the wall of large intrahepatic bile ducts (the so-called peribiliary glands). These EpCAMpos cell niches have been proven to harbor stem/progenitor cells with great importance in liver and biliary tree regeneration and in the pathophysiology of human diseases. The EpCAMpos progenitor cells within bile ductules are engaged in driving regenerative processes in chronic diseases affecting hepatocytes or interlobular bile ducts. The EpCAMpos population within peribiliary glands is activated when regenerative needs are finalized to repair large intra- or extra-hepatic bile ducts affected by chronic pathologies, including primary sclerosing cholangitis and ischemia-induced cholangiopathies after orthotopic liver transplantation. Finally, the presence of distinct EpCAMpos cell populations may explain the histological and molecular heterogeneity characterizing cholangiocarcinoma, based on the concept of multiple candidate cells of origin. This review aimed to describe the precise anatomical distribution of EpCAMpos populations within the liver and the biliary tree and to discuss their contribution in the pathophysiology of human liver diseases, as well as their potential role in regenerative medicine of the liver.

Extrahepatic bile ducts are characterized by the presence of peribiliary glands (PBGs), which represent stem cell niches implicated in biliary regeneration. Orthotopic liver transplantation may be complicated by non-anastomotic strictures (NAS) of the bile ducts, which have been associated with ischemic injury of PBGs and occur more frequently in livers obtained from donors after circulatory death than in those from brain-dead donors. The aims of the present study were to investigate the PBG phenotype in bile ducts after transplantation, the integrity of the peribiliary vascular plexus (PVP) around PBGs, and the expression of vascular endothelial growth factor-A (VEGF-A) by PBGs. Transplanted ducts obtained from patients who underwent liver transplantation were studied (N=62). Controls included explanted bile duct samples not used for transplantation (N=10) with normal histology. Samples were processed for histology, immunohistochemistry and immunofluorescence. Surface epithelium is severely injured in transplanted ducts; PBGs are diffusely damaged, particularly in ducts obtained from circulatory-dead compared to brain-dead donors. PVP is reduced in transplanted compared to controls. PBGs in transplanted ducts contain more numerous progenitor and proliferating cells compared to controls, show higher positivity for VEGF-A compared to controls, and express VEGF receptor-2. In conclusion, PBGs and associated PVP are damaged in transplanted extrahepatic bile ducts; however, an activation of the PBG niche takes place and is characterized by proliferation and VEGF-A expression. This response could have a relevant role in reconstituting biliary epithelium and vascular plexus and could be implicated in the genesis of non-anastomotic strictures.

The liver is a frontline immune site specifically designed to check and detect potential pathogens from the bloodstream to maintain a general state of immune hyporesponsiveness. One of the main functions of the liver is the regulation of iron homeostasis. The liver detects changes in systemic iron requirements and can regulate its concentration. Pathological states lead to the dysregulation of iron homeostasis which, in turn, can promote infectious and inflammatory processes. In this context, hepatic viruses deviate hepatocytes’ iron metabolism in order to better replicate. Indeed, some viruses are able to alter the expression of iron-related proteins or exploit host receptors to enter inside host cells. Lactoferrin (Lf), a multifunctional iron-binding glycoprotein belonging to the innate immunity, is endowed with potent antiviral activity, mainly related to its ability to block viral entry into host cells by interacting with viral and/or cell surface receptors. Moreover, Lf can act as an iron scavenger by both direct iron-chelation or the modulation of the main iron-related proteins. In this review, the complex interplay between viral hepatitis, iron homeostasis, and inflammation as well as the role of Lf are outlined.

Non-alcoholic fatty liver disease (NAFLD) is a chronic liver disease characterized by lipid accumulation in hepatocytes in the absence of excessive alcohol consumption. The global prevalence of NAFLD is constantly increasing. NAFLD is a disease spectrum comprising distinct stages with different prognoses. Non-alcoholic steatohepatitis (NASH) is a progressive condition, characterized by liver inflammation and hepatocyte ballooning, with or without fibrosis. The natural history of NAFLD is negatively influenced by NASH onset and by the progression towards advanced fibrosis. Pathogenetic mechanisms and cellular interactions leading to NASH and fibrosis involve hepatocytes, liver macrophages, myofibroblast cell subpopulations, and the resident progenitor cell niche. These cells are implied in the regenerative trajectories following liver injury, and impairment or perturbation of these mechanisms could lead to NASH and fibrosis. Recent evidence underlines the contribution of extra-hepatic organs/tissues (e.g., gut, adipose tissue) in influencing NASH development by interacting with hepatic cells through various molecular pathways. The present review aims to summarize the role of hepatic parenchymal and non-parenchymal cells, their mutual influence, and the possible interactions with extra-hepatic tissues and organs in the pathogenesis of NAFLD.

Cholangiocarcinoma (CCA) is a highly aggressive malignancy that emerges from the biliary tree. There are three major classes of CCA—intrahepatic, hilar (perihilar), or distal (extrahepatic)—according to the location of tumor development. Although CCA tumors are mainly derived from biliary epithelia (i.e., cholangiocytes), CCA can be originated from other cells, such as hepatic progenitor cells and hepatocytes. This heterogeneity of CCA may be responsible for poor survival rates of patients, limited effects of chemotherapy and radiotherapy, and the lack of treatment options and novel therapies. Previous studies have identified a number of neuroendocrine mediators, such as hormones, neuropeptides, and neurotransmitters, as well as corresponding receptors. The mediator/receptor signaling pathways play a vital role in cholangiocyte proliferation, as well as CCA progression and metastases. Agonists or antagonists for candidate pathways may lead to the development of novel therapies for CCA patients. However, effects of mediators may differ between healthy or cancerous cholangiocytes, or between different subtypes of receptors. This review summarizes current understandings of neuroendocrine mediators and their functional roles in CCA.

Niches containing stem/progenitor cells are present in different anatomical locations along the human biliary tree and within liver acini. The most primitive stem/progenitors, biliary tree stem/progenitor cells (BTSCs), reside within peribiliary glands located throughout large extrahepatic and intrahepatic bile ducts. BTSCs are multipotent and can differentiate towards hepatic and pancreatic cell fates. These niches’ matrix chemistry and other characteristics are undefined. Canals of Hering (bile ductules) are found periportally and contain hepatic stem/progenitor cells (HpSCs), participating in the renewal of small intrahepatic bile ducts and being precursors to hepatocytes and cholangiocytes. The niches also contain precursors to hepatic stellate cells and endothelia, macrophages, and have a matrix chemistry rich in hyaluronans, minimally sulfated proteoglycans, fetal collagens, and laminin. The microenvironment furnishes key signals driving HpSC activation and differentiation. Newly discovered third niches are pericentral within hepatic acini, contain Axin2+ unipotent hepatocytic progenitors linked on their lateral borders to endothelia forming the central vein, and contribute to normal turnover of mature hepatocytes. Their relationship to the other stem/progenitors is undefined. Stem/progenitor niches have important implications in regenerative medicine for the liver and biliary tree and in pathogenic processes leading to diseases of these tissues.

Non-alcoholic fatty liver disease is one of the most important causes of liver-related morbidity in children. In non-alcoholic fatty liver disease, the activation of liver resident macrophage pool is a central event in the progression of liver injury. The aims of the present study were to evaluate the polarization of liver macrophages and the possible role of Wnt3a production by macrophages in hepatic progenitor cell response in the progression of pediatric non-alcoholic fatty liver disease. 32 children with biopsy-proven non-alcoholic fatty liver disease were included. 20 out of 32 patients were treated with docosahexaenoic acid for 18 months and biopsies at the baseline and after 18 months were included. Hepatic progenitor cell activation, macrophage subsets and Wnt/β-catenin pathway were evaluated by immunohistochemistry and immunofluorescence. Our results indicated that in pediatric non-alcoholic fatty liver disease, pro-inflammatory macrophages were the predominant subset. Macrophage polarization was correlated with Non-alcoholic fatty liver disease Activity Score, ductular reaction, and portal fibrosis; docosahexaenoic acid treatment determined a macrophage polarization towards an anti-inflammatory phenotype in correlation with the reduction of serum inflammatory cytokines, with increased macrophage apoptosis, and with the up-regulation of macrophage Wnt3a expression; macrophage Wnt3a expression was correlated with β-catenin phosphorylation in hepatic progenitor cells and signs of commitment towards hepatocyte fate. In conclusion, macrophage polarization seems to have a key role in the progression of pediatric non-alcoholic fatty liver disease; the modulation of macrophage polarization could drive hepatic progenitor cell response by Wnt3a production.

INTRODUCTION:Nonalcoholic fatty liver disease (NAFLD) is one of the most important causes of liver-related morbidity and mortality in children. Recently, we have reported the effects of docosahexaenoic acid (DHA), the major dietary long-chain polyunsaturated fatty acids, in children with NAFLD. DHA exerts a potent anti-inflammatory activity through the G protein-coupled receptor (GPR)120. Our aim was to investigate in pediatric NAFLD the mechanisms underlying the effects of DHA administration on histo-pathological aspects, GPR120 expression, hepatic progenitor cell activation and macrophage pool. PATIENTS AND METHODS:20 children with untreated NAFLD were included. Children were treated with DHA for 18 months. Liver biopsies before and after the treatment were analyzed. Hepatic progenitor cell activation, macrophage pool and GPR120 expression were evaluated and correlated with clinical and histo-pathological parameters. RESULTS:GPR120 was expressed by hepatocytes, liver macrophages, and hepatic progenitor cells. After DHA treatment, the following modifications were present: i) the improvement of histo-pathological parameters such as NAFLD activity score, ballooning, and steatosis; ii) the reduction of hepatic progenitor cell activation in correlation with histo-pathological parameters; iii) the reduction of the number of inflammatory macrophages; iv) the increase of GPR120 expression in hepatocytes; v) the reduction of serine-311-phosphorylated nuclear factor kappa B (NF-κB) nuclear translocation in hepatocytes and macrophages in correlation with serum inflammatory cytokines. CONCLUSIONS:DHA could modulate hepatic progenitor cell activation, hepatocyte survival and macrophage polarization through the interaction with GPR120 and NF-κB repression. In this scenario, the modulation of GPR120 exploits a novel crucial role in the regulation of the cell-to-cell cross-talk that drives inflammatory response, hepatic progenitor cell activation and hepatocyte survival.

Over the last years, cholangiocytes, the cells that line the biliary tree, have been considered an important object of study for their biological properties which involves bile formation, proliferation, injury repair, fibrosis and angiogenesis. Cholangiocyte proliferation occurs in all pathologic conditions of liver injury where it is associated with inflammation and regeneration. During these processes, biliary cells start to secrete different cytokines, growth factors, neuropeptides and hormones which represent potential mechanisms for cross talk with other liver cells. Several studies suggest that hormones, and in particular, sex hormones, play a fundamental role in the modulation of the growth of this compartment in the injured liver which functionally conditions the progression of liver disease. Understanding the mechanisms of action and the intracellular pathways of these compounds on cholangiocyte pathophysiology will provide new potential strategies for the management of chronic liver diseases. The purpose of this review is to summarize the recent findings on the role of sex hormones in cholangiocyte proliferation and biology.