Biodegradable polydioxanone stents: histologic and structural effects in an experimental model of tracheal stenosis.

OBJECTIVES: To evaluate the biologic impact of polydioxanone (PDO) stenting in an animal model of inflammatory tracheal stenosis (TS). Additionally, to compare these results with those obtained in the same model without a stent and after placing one PDO stent in a healthy trachea. METHODS: 40 adult NZ rabbits were distributed into 3 groups: Group A, 8 animals with a healthy trachea and a PDO stent; group B, 17 rabbits with a TS and no stent; and group C, 15 animals with TS and a PDO stent. Histopathological studies included Masson's trichrome staining for submucosal fibrosis and Safranin O to assess structural integrity of cartilage. Morphometric analyses were performed in the 3 groups. RESULTS: Stent placement was successful in every case. Histological studies did not show a significant increase in tracheal wall collagen area and cartilage structure was not modified in those rabbits with a PDO stent, even in a TS scenario. Stent implantation permitted recovery of normal tracheal lumen levels in the TS model. CONCLUSIONS: PDO stenting in the normal trachea and in a model of TS neither caused increase in the collagen matrix nor modification of the cartilaginous support. Additionally, radial force exhibited by PDO stents was effective in restoring normal tracheal lumen when placed in a stenotic lesion. These findings suggest that they may be safe and useful in the setting of an acquired TS.

Tryptophan Modulation in Cancer-Associated Cachexia Mouse Models.

Cancer cachexia is a multifactorial syndrome that interferes with treatment and reduces the quality of life and survival of patients. Currently, there is no effective treatment or biomarkers, and pathophysiology is not clear. Our group reported alterations on tryptophan metabolites in cachectic patients, so we aim to investigate the role of tryptophan using two cancer-associated cachexia syngeneic murine models, melanoma B16F10, and pancreatic adenocarcinoma that is KPC-based. Injected mice showed signs of cancer-associated cachexia as reduction in body weight and raised spleen weight, MCP1, and carbonilated proteins in plasma. CRP and Myostatin also increased in B16F10 mice. Skeletal muscle showed a decrease in quadriceps weight and cross-sectional area (especially in B16F10). Higher expression of atrophy genes, mainly Atrogin1, was also observed. Plasmatic tryptophan levels in B16F10 tumor-bearing mice decreased even at early steps of tumorigenesis. In KPC-injected mice, tryptophan fluctuated but were also reduced and in cachectic patients were significantly lower. Treatment with 1-methyl-tryptophan, an inhibitor of tryptophan degradation, in the murine models resulted in the restoration of plasmatic tryptophan levels and an improvement on splenomegaly and carbonilated proteins levels, while changes in plasmatic inflammatory markers were mild. After the treatment, CCR2 expression in monocytes diminished and lymphocytes, Tregs, and CD8+, were activated (seen by increased in CD127 and CD25 expression, respectively). These immune cell changes pointed to an improvement in systemic inflammation. While treatment with 1-MT did not show benefits in terms of muscle wasting and atrophy in our experimental setting, muscle functionality was not affected and central nuclei fibers appeared, being a feature of regeneration. Therefore, tryptophan metabolism pathway is a promising target for inflammation modulation in cancer-associated cachexia.

Histological and structural effects of biodegradable polydioxanone stents in the rabbit trachea.

OBJECTIVES: The aim of this study was to evaluate the potential biologic effects caused by the successive placement of biodegradable polydioxanone (PDO) stents in the rabbit trachea. PDO stents could eventually induce a fibroproliferative reaction in the submucosa that could be beneficial in the treatment of malacia due to an increase in its consistency without impairing the tracheal lumen. METHODS: Sixteen adult NZ rabbits were distributed into 3 groups with different survival times according to the number of stents placed: 1 stent (14 weeks), 2 stents (28 weeks) and 3 stents (42 weeks). Stent insertion was performed endoscopically in the cervical trachea of the animal. Histopathological studies included Masson's trichrome staining for submucosal fibrosis and Safranin O to assess the structural integrity of cartilage. Potential inflammatory changes were analysed by means of immunohistochemistry determining the number of CD45-positive cells. RESULTS: Stent placement was successful in every case. Histological studies did not show a statistically significant increase in tracheal wall collagen area and cartilage structure was not modified in those rabbits with 1 or more PDO stents inserted compared to non-stented tracheal sections. Furthermore, no statistically significant changes in the number of CD45+ cells were observed in stented tracheal segments compared to normal tracheal tissues. CONCLUSIONS: According to our data, successive PDO stenting caused mild inflammatory changes in the tracheal wall and no increase in the collagen matrix, and the cartilaginous support was not modified during a long follow-up period (up to 42 weeks). These findings suggest that they may be safe and show good biocompatibility in the long term.

Implementation of a peroral endoscopic myotomy program.

INTRODUCTION: the aim of our study was to develop a peroral endoscopic myotomy (POEM) program in our Unit following a two-step sequence: training on animal models and supervision by an experienced endoscopist during the first human cases. METHODS: a single endoscopist experienced in advanced endoscopy was trained in POEM. After observing POEM in referral centers, training was implemented on swine models (preclinical phase). Technical aspects and adverse events were prospectively recorded. A first subset of cases (group A) was compared to a second one (group B) to assess our progression. Finally, POEM was implemented in humans under the supervision of an experienced endoscopist (clinical phase). The outcomes and adverse events were prospectively recorded. RESULTS: during the preclinical phase, 15 POEM procedures were performed on live pigs. Severe adverse events (AE) were less frequent in group B than in group A (12 % vs 57 %, p = 0.07). After nine cases, a plateau of adverse events was reached. During the clinical phase, eleven POEM procedures were performed in patients under expert supervision. Technical and clinical (Eckardt score ≤ 3) success were 100 % and 91 %, respectively (follow-up 3-21 months). In two cases, intervention of an experienced endoscopist was required (cases 2 and 3) because of a difficult orientation at the esophagogastric junction. One mild pneumoperitoneum occurred, with no severe adverse events reported. CONCLUSIONS: training in animal models and supervision by an experienced endoscopist during the first cases could provide the necessary skills to perform POEM safely and effectively.

Omentectomy Prevents Metabolic Syndrome By Reducing Appetite and Body Weight In A Diet-Induced Obesity Rat Model.

Visceral fat deposition is associated with impairment of glucose and lipid metabolism while leptin levels are frequently related to subcutaneous fat area. At present, there is considerable controversy regarding the role of visceral adipose tissue accumulation in the development of metabolic syndrome (MS). Here we show the effects of omentectomy on the liver and MS in a diet induced obesity rat model. Our results reveal that undergoing omentectomy previously the establishment of the diet-induced-obesity reduced significantly body weight gain and avoid the development of MS, including non-alcoholic fatty liver disease. Intriguingly, the significantly lower body weight gain was due to decreased food intake. Omentum drives obesity progression through leptin resistance mediated by C-reactive protein, Interleucin (IL)-6 and high lipolysis activity. Omentum removal reversed immediately the increased plasma levels of CRP and IL-6 and gradually food intake, weight gain, and features of MS in diet-induced-obesity. Omentectomy caused no changes in normal-weigh-rats. This report displays causal mechanism by which omentum promotes obesity and propose omentectomy as a promising procedure in MS prevention.

Vectorization of ultrasound-responsive nanoparticles in placental mesenchymal stem cells for cancer therapy.

A new platform constituted by engineered responsive nanoparticles transported by human mesenchymal stem cells is here presented as a proof of concept. Ultrasound-responsive mesoporous silica nanoparticles are coated with polyethylenimine to favor their effective uptake by decidua-derived mesenchymal stem cells. The responsive-release ability of the designed nanoparticles is confirmed, both in vial and in vivo. In addition, this capability is maintained inside the cells used as carriers. The migration capacity of the nanoparticle-cell platform towards mammary tumors is assessed in vitro. The efficacy of this platform for anticancer therapy is shown against mammary tumor cells by inducing the release of doxorubicin only when the cell vehicles are exposed to ultrasound.

TGF-ß antagonist attenuates fibrosis but not luminal narrowing in experimental tracheal stenosis.

OBJECTIVE: Acquired tracheal stenosis (ATS) is an unusual disease often secondary to prolonged mechanical trauma. Acquired tracheal stenosis pathogenesis involves inflammation and subsequent fibrosis with narrowing of the tracheal lumen. Transforming growth factor-ß1 (TGF-ß) represents a pivotal factor in most fibrotic processes, and therefore a potential target in this context. The aim of this study is to analyze the role of TGF-ß as a target for anti-fibrotic interventions in tracheal stenosis. METHODS: Human stenotic tracheobronchial tissues from patients with benign airway stenosis and normal controls from pneumonectomy specimens were analyzed. Tracheal stenosis was induced in adult NZ rabbits by a circumferential thermal injury to the mucosa during open surgery and re-anastomosis. Rabbits were treated postoperatively with a peritracheal collagen sponge containing a TGF-ß peptide antagonist (p17) or vehicle. Fibrosis was determined by Masson's trichrome staining, and smooth muscle cell α-actin+ (α-SMA+ Confirm accuracy.) myofibroblasts, connective tissue growth factor (CTGF), and p-Smad2/3 expression by immunohistochemistry. RESULTS: Human and rabbit stenotic tissues showed extensive submucosal fibrosis, characterized by significantly increased α-SMA+ myofibroblasts and CTGF expression. In human stenotic lesions, increased p-Smad2/3+ nuclei were also observed. p17 treatment significantly reduced the fibrotic thickness, as well as the density of α-SMA+ myofibroblasts and CTGF+ cells in rabbit stenotic lesions, but failed to improve the luminal area. CONCLUSION: ATS is characterized by a TGF-ß dependent fibrotic process, but reduction of the fibrotic component by TGF-ß1 antagonist therapy was not sufficient to improve tracheal narrowing, suggesting that fibrosis may not be the main contributor to luminal stenosis. LEVEL OF EVIDENCE: NA. Laryngoscope, 127:561-567, 2017.

NADPH oxidase is implicated in the pathogenesis of oxidative phosphorylation dysfunction in mice fed a high-fat diet.

The aim of this study was to evaluate the role of NADPH oxidase (NADPHox) in the pathogenesis of oxidative phosphorylation (OXPHOS) dysfunction as found in mice fed a high-fat diet (HFD). C57BL/6J mice were distributed in four groups: WT/SCD: six wild-type (WT) mice fed a standard chow diet (SCD); WT/HFD, six WT mice fed a HFD; NOX2(-/-)/SCD, six NADPHox-deficient mice on a SCD; (4) NOX2(-/-)/HFD, six NADPHox-deficient mice on a HFD. After 32 weeks, we studied the liver for: histology; OXPHOS complex activity; fully assembled OXPHOS complexes and their subunits; gene expression of OXPHOS subunits; oxidative and nitrosative stress; and oxidative DNA damage. In the liver of WT/HFD mice, we found a significant decreased in the activity of all OXPHOS complexes, in fully assembled complexes, in the amount of OXPHOS subunits, and in gene expression of mitochondrial DNA-encoded subunits. 8-hydroxy-2'-deoxyguanosine was only increased in mitochondrial DNA. The liver of NOX(-/-)/HFD mice showed mild steatosis but no non-alcoholic steatohepatitis (NASH) lesions were found. OXPHOS activity, OXPHOS subunits, and assembly of subunits into OXPHOS complexes were normal in these mice. We conclude that this study shows that NADPH deficiency protects mice from developing OXPHOS dysfunction and NASH caused by a HFD.

Blockade of cell adhesion molecules enhances cell engraftment in a murine model of liver cell transplantation.

AIM: OLT is the best alternative for patients with end-stage liver diseases. However, as the need for organs surpasses donor availability, alternatives to OLT are required. LCT could be a useful option versus OLT in several patients even though its low cell-engraftment hampers its efficiency. Endothelial cell barrier is the main obstacle for the implantation of cells into the parenchyma. Our study has focused on the modification of the endothelial barrier with monoclonal antibodies against adhesion molecules in order to increase cell engraftment in a mouse model of liver cell transplantation. METHODS: Anti-mouse CD54 and anti-mouse CD61 antibodies were administered intrasplenically to healthy mice within 60 min prior to stem cell transplantation. Animals were sacrificed either short term at 2h or middle term seven days after transplantation. Immunohistochemical techniques to detect alkaline phosphatase activity were used to identify the transplanted cells within the liver parenchyma. RESULTS: Anti-CD54 and anti-CD61 administration increases vascular patency and cell engraftment. This represents a 32% and 45% increase, respectively, of engrafted cells compared to the control (p<0.05). CONCLUSION: Modification of the vascular wall with monoclonal antibodies against endothelial adhesion molecules before cell transplantation enhances cell engraftment into the mouse liver.

Ependymal cell differentiation, from monociliated to multiciliated cells.

Primary and motile cilia differ in their structure, composition, and function. In the brain, primary cilia are immotile signalling organelles present on neural stem cells and neurons. Multiple motile cilia are found on the surface of ependymal cells in all brain ventricles, where they contribute to the flow of cerebrospinal fluid. During development, monociliated ependymal progenitor cells differentiate into multiciliated ependymal cells, thus providing a simple system for studying the transition between these two stages. In this chapter, we provide protocols for immunofluorescence staining of developing ependymal cells in vivo, on whole mounts of lateral ventricle walls, and in vitro, on cultured ependymal cells. We also provide a list of markers we currently use to stain both types of cilia, including proteins at the ciliary membrane and tubulin posttranslational modifications of the axoneme.

High-fat diet decreases activity of the oxidative phosphorylation complexes and causes nonalcoholic steatohepatitis in mice.

Nonalcoholic fatty liver disease (NAFLD) is the most frequent histological finding in individuals with abnormal liver-function tests in the Western countries. In previous studies, we have shown that oxidative phosphorylation (OXPHOS) is decreased in individuals with NAFLD, but the cause of this mitochondrial dysfunction remains uncertain. The aims of this study were to determine whether feeding mice a high-fat diet (HFD) induces any change in the activity of OXPHOS, and to investigate the mechanisms involved in the pathogenesis of this defect. To that end, 30 mice were distributed between five groups: control mice fed a standard diet, and mice on a HFD and treated with saline solution, melatonin (an antioxidant), MnTBAP (a superoxide dismutase analog) or uric acid (a scavenger of peroxynitrite) for 28 weeks intraperitoneously. In the liver of these mice, we studied histology, activity and assembly of OXPHOS complexes, levels of subunits of these complexes, gene expression of these subunits, oxidative and nitrosative stress, and oxidative DNA damage. In HFD-fed mice, we found nonalcoholic steatohepatitis, increased gene expression of TNFα, IFNγ, MCP-1, caspase-3, TGFß1 and collagen α1(I), and increased levels of 3-tyrosine nitrated proteins. The activity and assembly of all OXPHOS complexes was decreased to about 50-60%. The amount of all studied OXPHOS subunits was markedly decreased, particularly the mitochondrial-DNA-encoded subunits. Gene expression of mitochondrial-DNA-encoded subunits was decreased to about 60% of control. There was oxidative damage to mitochondrial DNA but not to genomic DNA. Treatment of HFD-fed mice with melatonin, MnTBAP or uric acid prevented all changes observed in untreated HFD-fed mice. We conclude that a HFD decreased OXPHOS enzymatic activity owing to a decreased amount of fully assembled complexes caused by a reduced synthesis of their subunits. Antioxidants and antiperoxynitrites prevented all of these changes, suggesting that nitro-oxidative stress played a key role in the pathogenesis of these alterations. Treatment with these agents might prevent the development of NAFLD in humans.

Sp1 and Sp3 transcription factors mediate leptin-induced collagen α1(I) gene expression in primary culture of male rat hepatic stellate cells.

Mechanisms by which leptin stimulates collagen α(1)(I) [Col1a(I)] gene expression are unclear. The purposes of this study were to identify the trans-acting factors and cis-acting elements in Col1a(I) promoter involved in this effect as well as the pathways that are implicated. In primary cultures of rat hepatic stellate cells (HSCs), we measured the effects of leptin on Col1a(I) gene and protein expression and on the binding of nuclear proteins to the Col1a(I) promoter. We found that leptin increased Col1a(I) gene and protein expression in activated HSCs. Transient transfections showed that leptin exerted its effects through elements located between -220 and -112 bp of the Col1a(I) promoter. Gel retardation assays demonstrated that leptin induced the binding of transcription factors specific protein (Sp)-1 and Sp3 to two elements located between -161 and -110 bp of the Col1a(I) promoter. Leptin-induced Sp1/Sp3 phosphorylation, but this effect was suppressed by inhibiting or silencing Janus kinase-2, phosphatidylinositol-3-kinase, nonphagocytic adenine dinucleotide phosphate (NADPH) oxidase, or ERK1/2, by the use of antioxidants or catalase, or by preventing protein-aldehyde adduct formation. Leptin provoked oxidative stress, aldehyde-protein adduct formation, and increased gene expression of some components of the NADPH oxidase complex. In conclusion, in HSCs, leptin up-regulates Col1a(I) gene expression after activating NADPH oxidase, inducing oxidative stress, aldehyde-protein adduct formation, and ERK1/2 phosphorylation, which in turn activates Sp1/Sp3 and provokes the binding of these two factors to regulatory elements located between -161 and -110 bp of the Col1a(I) promoter. These findings may contribute to a better understanding of mechanisms involved in the leptin-induced liver fibrosis.

Blockade of endothelial G(i) protein enhances early engraftment in intraportal cell transplant to mouse liver.

The limited availability of liver donors and recent progress in cell therapy technologies has centered interest on cell transplantation as a therapeutic alternative to orthotopic liver transplant for restoring liver function. Following transplant by intraportal perfusion, the main obstacle to cell integration in the parenchyma is the endothelial barrier. Transplanted cells form emboli in the portal branches, inducing ischemia and reperfusion injury, which cause disruption of endothelial impermeability and activate the immune system. Approximately 95% of transplanted cells fail to implant and die within hours by anoikis or are destroyed by the host immune system. Intravascular perfusion of Bordetella pertussis toxin (PTx) blocks endothelial G(i) proteins and acts as a reversible inducer of actin cytoskeleton reorganization, leading to interruption of cell confluence in vitro and increased vascular permeability in vivo. PTx treatment of the murine portal vascular tree 2 h before intraportal perfusion of embryonic stem cells facilitated rapid cell engraftment. By 2 h postperfusion, the number of implanted cells in treated mice was more than fivefold greater than in untreated controls, a difference that was maintained to at least 30 days posttransplant. We conclude that prior to cell transplant, PTx blockade of the G(i) protein pathway in liver endothelium promotes rapid, efficient cell implantation in liver parenchyma, and blocks chemokine receptor signaling, an essential step in early activation of the immune system.

Hepcidin treatment in Hfe-/- mice diminishes plasma iron without affecting erythropoiesis.

BACKGROUND: Iron is essential for mammalian metabolism and its cellular concentration is controlled by regulating its acquisition and storage. Haemochromatosis is a condition involving iron overload that is characterised by increased duodenal iron absorption and a progressive accumulation of iron in vital organs. Hepcidin is the main hormone that regulates iron homoestasis and it is secreted by the liver. MATERIALS AND METHODS: We have studied how extended hepcidin administration affects the iron load status, plasma and tissue iron concentration, erythropoiesis and the expression of proteins involved on iron homeostasis in haemochromatotic (Hfe(-/-)) and wild-type mice. RESULTS: Hepcidin reverted the high plasma iron concentrations in Hfe(-/-) mice to normal values. The high concentration of hepatic iron was not altered in the liver of these Hfe(-/-) mice. Hepcidin administration did not disturb erythropoiesis in either Hfe(-/-) or wild-type mice and likewise, hepcidin did not modify the expression of any protein analysed in the liver, duodenum or spleen of Hfe(-/-) and wild-type mice. These data confirm that hepcidin administration diminishes plasma iron concentrations. CONCLUSION: Treatment with sustained doses of hepcidin diminishes plasma iron concentrations in Hfe(-/-) mice.

In vivo effect of an luteinizing hormone-releasing hormone analog on vascular endothelial growth factor and epidermal growth factor receptor expression in mammary tumors.

BACKGROUND: The hypothalamic luteinizing hormone-releasing hormone (LHRH) is well known for its role in the control of pituitary gonadotropin secretion and it has demonstrated a direct antiproliferative effect on some cancer cell lines of LHRH and its synthetic analogs. The study was designed to assess whether administration of the LHRH analog (goserelin) has any effect on the expression of the vascular endothelial growth factor (VEGF) and the epidermal growth factor receptor (EGFR) in rats with N-nitroso-N-methylurea (NMU)-induced-mammary tumors " in vivo". MATERIALS AND METHODS: The animals with tumors were assessed after acute or chronic treatment with goserelin, and in all the animals VEGF and EGFR expression was examined both in plasma and tumor homogenates by enzyme immunoassay. RESULTS: The basal plasma values of VEGF were lower in the healthy control group than in rats with NMU-induced tumors ( P = 0.025). Following acute treatment with goserelin, VEGF expression in plasma increased above basal levels after 60 min ( P = 0.05) and dropped during chronic treatment. Likewise, in the tumor homogenate the mean VEGF expression was higher at 60 min post-goserelin administration than the basal levels, although VEGF expression then diminished at 90 min. Plasma EGFR expression was higher in rats with NMU-induced tumors than in healthy controls ( P Conclusions: The results allow us to conclude that goserelin may exert a short-term stimulatory effect on the release of VEGF, as well as a long-term inhibitory effect on VEGF but not EGFR expression.

Bone marrow transplantation into hemochromatotic mice decreases hepatic and duodenal iron overload.

Human hereditary hemochromatosis is a disorder of iron homeostasis characterized by increased absorption of iron and its deposition in parenchymal organs. The maintenance of iron homeostasis is regulated by molecules involved in the absorption, transport, storage and redox of iron. The potential of hematopoietic stem cell therapy for liver diseases has been studied in some experimental animal models. Our objective was to evaluate the effect of bone marrow transplantation from wild type mice on the status of iron overload in Hfe knockout hemochromatotic mice (Hfe(-/-)). The transplanted cells were detected in the liver (11% of the total cells) and characterized as hepatocytes and myofibroblasts. They were also detected in the duodenum and characterized as myofibroblasts. The iron content in the Hfe(-/-) mice descended 2.9-fold in the liver and 2.4-fold in the duodenum 6 months after transplantation. Non-significant changes of relative mRNA abundance of genes of iron metabolism were observed in the liver and duodenum of Hfe(-/-) transplanted mice. At 6 months after transplantation the proportion of Hfe mRNA in Hfe(-/-) mice reached 3.8% of the levels in wild type mice in the liver and 1.6% in the duodenum. In conclusion, adult stem cells from bone marrow transplant were able to differentiate into hepatocytes and myofibroblasts in hemochromatotic mice. Bone marrow transplantation assisted in reducing the iron overload in this murine model of hemochromatosis. These findings might contribute to the knowledge of pathways involved in the regulatory system of iron homeostasis.

Increased mitochondrial respiratory chain enzyme activities correlate with minor extent of liver damage in mice suffering from erythropoietic protoporphyria.

Mitochondrial dysfunction might play a role in the pathogenesis of liver damage in erythropoietic protoporphyria (EPP). Changes in mitochondrial respiratory chain activities were evaluated in the Fech(m1pas)/Fech(m1pas) mouse model for EPP. Mice from different strains congenic for the same ferrochelatase germline mutation manifest variable degrees of hepatobiliary injury. Protoporphyric animals bred into the C57BL/6J background showed a higher degree of hepatomegaly and liver damage as well as higher protoporphyrin (PP) accumulation than those bred into the SJL/J and BALB/cJ backgrounds. Whereas mitochondrial respiratory chain activities remained unchanged in the liver of protoporphyric mice C57BL/6J, they were increased in protoporphyric mice from both SJL/J and BALB/cJ backgrounds, when compared to wild-type animals. Mitochondrial respiratory chain activities were increased in Hep G2 cell line after accumulation of PP following addition of aminolevulinic acid. As a direct effect of these elevated mitochondrial activities, in both hepatic cells from mutant mouse strains and Hep G2 cells, adenosine 5'-triphosphate (ATP) levels significantly increased as the intracellular PP concentration was reduced. These results indicate that PP modifies intracellular ATP requirements as well as hepatic mitochondrial respiratory chain enzymatic activities and further suggest that an increase of these activities may provide a certain degree of protection against liver damage in protoporphyric mice.

Sialoadenectomy alters liver cell turn-over and function in mice.

In rodents, submandibular salivary glands accumulate a number of biologically active peptides, and release some of them to both saliva and the bloodstream. Surgical removal of these glands (sialoadenectomy) alters the ability of the liver to regenerate after partial hepatectomy. We show here that 5 weeks after surgery, the liver of sialoadenectomized mice contained 40% fewer hepatocytes than the liver of sham-operated mice. We did not obtain evidence of necrotic cell death after surgery. In contrast, sialoadenectomy transiently increased apoptotic hepatocyte death, as revealed by terminal deoxynucleotidyl transferase(TdT)-mediated dUTP nick-end labeling (TUNEL) assay. DNA synthesis was determined in vivo by the incorporation of bromo-deoxyuridine (BrdU) into hepatocyte nuclei. BrdU-labeling progressively increased after sialoadenectomy. We conclude that sialoadenectomy induced a transient wave of apoptotic cell death followed by a rise in DNA synthesis but not by cell division. This reduced cell number but increased mean cell volume. In spite of these alterations in cellularity, the liver responded adequately to several stressful conditions, as judged by the lack of any differential effect of sialoadenectomy on liver glycogen and plasma glucose concentration after immobilization, aggressive encounter, or fasting. However, the liver of sialoadenectomized mice was more sensitive to the effect of a non-lethal dose of bacterial lipopolysaccharide (LPS) combined with d-galactosamine, as shown by the enhanced rise in plasma alanine aminotransferase and aspartate aminotransferase, and liver myeloperoxidase (MPO) activities. All these results indicate that a submandibular salivary glands-liver axis is involved in the maintenance of liver structure in mice. A disturbance of this axis induces an adaptive response that preserves the metabolic function of the liver but renders it more sensitive to bacterial endotoxins.