Lipidomic Approaches to Study HDL Metabolism in Patients with Central Obesity Diagnosed with Metabolic Syndrome.

The metabolic syndrome (MetS) is a cluster of cardiovascular risk factors characterised by central obesity, atherogenic dyslipidaemia, and changes in the circulating lipidome; the underlying mechanisms that lead to this lipid remodelling have only been partially elucidated. This study used an integrated "omics" approach (untargeted whole serum lipidomics, targeted proteomics, and lipoprotein lipidomics) to study lipoprotein remodelling and HDL composition in subjects with central obesity diagnosed with MetS (vs. controls). Compared with healthy subjects, MetS patients showed higher free fatty acids, diglycerides, phosphatidylcholines, and triglycerides, particularly those enriched in products of de novo lipogenesis. On the other hand, the "lysophosphatidylcholines to phosphatidylcholines" and "cholesteryl ester to free cholesterol" ratios were reduced, pointing to a lower activity of lecithin cholesterol acyltransferase (LCAT) in MetS; LCAT activity (directly measured and predicted by lipidomic ratios) was positively correlated with high-density lipoprotein cholesterol (HDL-C) and negatively correlated with body mass index (BMI) and insulin resistance. Moreover, many phosphatidylcholines and sphingomyelins were significantly lower in the HDL of MetS patients and strongly correlated with BMI and clinical metabolic parameters. These results suggest that MetS is associated with an impairment of phospholipid metabolism in HDL, partially led by LCAT, and associated with obesity and underlying insulin resistance. This study proposes a candidate strategy to use integrated "omics" approaches to gain mechanistic insights into lipoprotein remodelling, thus deepening the knowledge regarding the molecular basis of the association between MetS and atherosclerosis.

The Long-Term Impact of Renin-Angiotensin System (RAS) Inhibition on Cardiorenal Outcomes (LIRICO): A Randomized, Controlled Trial.

BACKGROUND: The comparative effectiveness of treatment with angiotensin-converting enzyme (ACE) inhibitors, angiotensin receptor blockers (ARBs), or their combination in people with albuminuria and cardiovascular risk factors is unclear. METHODS: In a multicenter, randomized, open label, blinded end point trial, we evaluated the effectiveness on cardiovascular events of ACE or ARB monotherapy or combination therapy, targeting BP<130/80 in patients with moderate or severe albuminuria and diabetes or other cardiovascular risk factors. End points included a primary composite of cardiovascular death, nonfatal myocardial infarction, nonfatal stroke, and hospitalization for cardiovascular causes and a revised end point of all-cause mortality. Additional end points included ESRD, doubling of serum creatinine, albuminuria, eGFR, BP, and adverse events. RESULTS: Because of slow enrollment, the trial was modified and stopped 41% short of targeted enrollment of 2100 participants, corresponding to 35% power to detect a 25% reduced risk in the primary outcome. Our analysis included 1243 adults, with median follow-up of 2.7 years. Efficacy outcomes were similar between groups (ACE inhibitor versus ARB, ACE inhibitor versus combination, ARB versus combination) as were rates of serious adverse events. The rate of permanent discontinuation for ARB monotherapy (6.3%) was significantly lower than for ACE inhibitor monotherapy (15.7%) or combined therapy (18.3%). CONCLUSIONS: Patients may tolerate ARB monotherapy better than ACE inhibitor monotherapy. However, data from this trial and similar trials, although as yet inconclusive, show no trend suggesting differences in mortality and renal outcomes with ACE inhibitors or ARBs as dual or monotherapy in patients with albuminuria and diabetes or other cardiovascular risk factors.

Genes and miRNA expression signatures in peripheral blood mononuclear cells in healthy subjects and patients with metabolic syndrome after acute intake of extra virgin olive oil.

Extra virgin olive oil (EVOO) consumption has been associated with reduced cardiovascular risk but molecular mechanisms underlying its beneficial effects are not fully understood. Here we aimed to identify genes and miRNAs expression changes mediated by acute high- and low-polyphenols EVOO intake. Pre- and post-challenge gene and miRNAs expression analysis was performed on the peripheral blood mononuclear cells (PBMCs) of 12 healthy subjects and 12 patients with metabolic syndrome (MS) by using microarray and RT-qPCR. In healthy subjects, acute intake of EVOO rich in polyphenols was able to ameliorate glycaemia and insulin sensitivity, and to modulate the transcription of genes and miRNAs involved in metabolism, inflammation and cancer, switching PBMCs to a less deleterious inflammatory phenotype; weaker effects were observed in patients with MS as well as in healthy subjects following low-polyphenol EVOO challenge. Concluding, our study shows that acute high-polyphenols EVOO intake is able to modify the transcriptome of PBMCs through the modulation of different pathways associated with the pathophysiology of cardio-metabolic disease and cancer. These beneficial effects are maximized in healthy subjects, and by the use of EVOO cultivars rich in polyphenols. Nutrigenomic changes induced by EVOO thus legitimate the well-known beneficial effects of EVOO in promoting human health and, potentially, preventing the onset of cardiovascular disease and cancer.

A novel cluster of patients with Familial Mediterranean Fever (FMF) in southern Italy.

BACKGROUND: Familial Mediterranean Fever (FMF) is an autosomal recessive autoinflammatory disorder characterised by recurrent attacks of fever and serositis (peritonitis, pleuritic or synovitis) affecting mainly populations of Mediterranean origin. AIM: To describe a relatively new cluster of FMF subjects from Apulia and Basilicata regions (southern Italy). PATIENTS AND METHODS: Subjects were screened for FMF using the Tel-Hashomer criteria and genetic analysis. Demographic data were taken from patients' files and direct interviews. Patients were investigated about attack duration, intensity and site, body temperature, skin manifestations and overall quality of life before and after treatment with colchicine. Inflammatory parameters were also measured between these periods. RESULTS: Forty-nine subjects had FMF (M : F = 26 : 23, age 38 years ± 2 SE) and followed-up up to 8 years. The age at disease onset was 22·1 years ± 1·2SE and the diagnostic delay was 15·5 years ± 1·9SE. The majority of patients (82%) suffered from abdominal pain, and 35% had undergone prior abdominal surgery or laparotomy. Severity score (ISSF) was mild in 43% of patients and intermediate in 57% of patients. Serum amyloid A (SAA) was increased in 20% of patients (16·9 ± 3·7, normal range < 6·4 mg/dL). In over 95% of patients, inflammation markers, duration and intensity of febrile painful attacks, quality of life and ISSF score improved dramatically following colchicine treatment. CONCLUSION: The Apulia region represents a new endemic area for FMF. Clinical presentation of FMF can be misleading and requires a complete and early workup to recognise the disease and avoid unjustified surgery. Colchicine remains the gold standard therapy to prevent FMF attacks and fatal long-term complications.

Short-term benefits of an unrestricted-calorie traditional Mediterranean diet, modified with a reduced consumption of carbohydrates at evening, in overweight-obese patients.

The Mediterranean diet (MeD) is believed to promote health; nevertheless, changes in the nutritional patterns in the Mediterranean area (increased intake of refined carbohydrates/saturated fats; reduced fibers intake; main calorie load shifted to dinner) led to reduced MeD benefits in recent decades. We retrospectively investigated the effects of a MeD with a low intake of refined carbohydrates in the evening ("MeDLowC") on body weight (BW) and metabolic profile of overweight/obese subjects. According to their adherence to MeDLowC, subjects were classified into 44 (41%) individuals with "excellent" adherence and 63 (59%) with "poor" adherence. Nutritional counseling induced an improvement in BW, glucose metabolism and liver transaminases in both groups, with an increased magnitude of these effects in the "Excellent" adherence group. "Excellent" adherence to MeDLowC improved insulin sensitivity and lipid metabolism. In conclusion, MeD with a restriction of carbohydrates in the evening significantly ameliorates obesity and associated metabolic complications.

The p66Shc protein controls redox signaling and oxidation-dependent DNA damage in human liver cells.

The p66Shc protein mediates oxidative stress-related injury in multiple tissues. Steatohepatitis is characterized by enhanced oxidative stress-mediated cell damage. The role of p66Shc in redox signaling was investigated in human liver cells and alcoholic steatohepatitis. HepG2 cells with overexpression of wild-type or mutant p66Shc, with Ser36 replacement by Ala, were obtained through infection with recombinant adenoviruses. Reactive oxygen species and oxidation-dependent DNA damage were assessed by measuring dihydroethidium oxidation and 8-hydroxy-2'-deoxyguanosine accumulation into DNA, respectively. mRNA and protein levels of signaling intermediates were evaluated in HepG2 cells and liver biopsies from control and alcoholic steatohepatitis subjects. Exposure to H2O2 increased reactive oxygen species and phosphorylation of p66Shc on Ser36 in HepG2 cells. Overexpression of p66Shc promoted reactive oxygen species synthesis and oxidation-dependent DNA damage, which were further enhanced by H2O2. p66Shc activation also resulted in increased Erk-1/2, Akt, and FoxO3a phosphorylation. Blocking of Erk-1/2 activation inhibited p66Shc phosphorylation on Ser36. Increased p66Shc expression was associated with reduced mRNA levels of antioxidant molecules, such as NF-E2-related factor 2 and its target genes. In contrast, overexpression of the phosphorylation defective p66Shc Ala36 mutant inhibited p66Shc signaling, enhanced antioxidant genes, and suppressed reactive oxygen species and oxidation-dependent DNA damage. Increased p66Shc protein levels and Akt phosphorylation were observed in liver biopsies from alcoholic steatohepatitis compared with control subjects. In human alcoholic steatohepatitis, increased hepatocyte p66Shc protein levels may enhance susceptibility to DNA damage by oxidative stress by promoting reactive oxygen species synthesis and repressing antioxidant pathways.

Contrast-enhanced ultrasound in the diagnosis of hepatocellular carcinoma.

PURPOSE: The American Association for the Study of Liver Diseases and the European Association for the Study of the Liver exclude any role of contrast-enhanced ultrasound (CEUS) in the diagnosis of hepatocellular carcinoma (HCC) while the Italian Association for the Study of the Liver suggests its use for larger HCC. This study evaluated the accuracy of CEUS in comparison with computed tomography (CT) in the diagnosis of HCC and of residual of HCC after treatment. MATERIALS AND METHODS: We retrospectively evaluated 124 patients with 148 HCC nodules: 34 small (≤20 mm) and 114 large nodules (>20 mm). Ninety-three patients underwent treatment [one resection, 23 transcatheter arterial chemoembolisation (TACE), 37 radiofrequency ablation (RFA), 32 TACE/RFA combined with sorafenib]. The diagnosis of HCC on CEUS was confirmed by the typical pattern of arterial enhancement and portal and/or venous phase washout. RESULTS: We performed 90 CEUS for the initial diagnosis of HCC in 85 patients and 107 CEUS for the diagnosis of residual HCC after 1-month treatment in 92 patients. Sensitivity, specificity, positive predictive value, negative predictive value of CEUS and CT in the initial diagnosis of HCC were: 63 vs 92, 100 vs 100, 100 vs 100, 9 vs 25 for small HCC; 77 vs 92, 100 vs 100, 100 vs 100, 13 vs 22 for large HCC. In the diagnosis of residual of HCC, CEUS had a sensitivity of 70 % for small nodules and 76 % for large nodules, with an overall specificity of 100 %. CONCLUSION: CEUS is useful in the initial diagnosis and in the assessment of necrosis after RFA and TACE of HCC nodules.

Moving towards personalised therapy in patients with hepatocellular carcinoma: the role of the microenvironment.

The goal of personalised therapy based on hepatocellular carcinoma (HCC) molecular characteristics is still beyond our grasp. Systemic treatments show poor efficacy, mainly because of the great heterogeneity of the tumour. Indeed, differences in aetiology, disease stage and biochemical composition of the fibrotic liver make cirrhosis itself a highly dyshomogeneous disease. Cancer cells grow in a cirrhotic microenvironment, interacting with stromal cells and engaging matrix components that differ from patient to patient, hampering the development of drugs to treat all patients. Growing evidence suggests a role for the cross-talk between HCC and the host stroma in driving disease progression and hence prognosis and survival. Many efforts have been devoted to identifying genes responsible for good or bad prognosis, but no study has yet proven helpful in guiding therapeutic choices and management over time, also taking into account the development of drug resistance. The questions of what to target and in which patient are still unsolved. In the personalised therapy scenario, the patient rather than the disease becomes the target of the therapy. However, this still requires an evidence-based medical approach. Herein, we will discuss how individual differences in terms of quality and quantity of the tissue microenvironment components affect progression of HCC. Then, we will highlight potential druggable pathways, also considering ongoing clinical trials. The development of biomarkers will be discussed in the light of new experimental research conducted with the aim of moving towards personalised therapy in patients with HCC.

Nuclear receptors expression chart in peripheral blood mononuclear cells identifies patients with Metabolic Syndrome.

BACKGROUND: Nuclear receptors are a class of 48 ligand-activated transcription factors identified as key players of metabolic and developmental processes. Most of these receptors are potential targets for pharmacological strategies in the Metabolic Syndrome. In the present study, we analyzed changes in the mRNA expression of nuclear receptors in the peripheral blood mononuclear cells of patients with Metabolic Syndrome, in order to identify novel biomarkers of disease and candidate targets for putative therapeutical approaches. METHODS AND RESULTS: We enrolled thirty healthy controls (14 M:16 F) and thirty naïve patients (16 M: 14 F; >3 criteria for Metabolic Syndrome upon Adult Treatment Panel III) without organ damage. Using quantitative real-time PCR, we assessed the expression patterns of nuclear receptors in peripheral blood mononuclear cells. 33/48 nuclear receptors were expressed in peripheral blood mononuclear cells. In patients with Metabolic Syndrome, we found a significant down-regulation of the entire PPAR, NR4A and RAR families, together with a repression of RXRα, VDR, and Rev-Erbα. Furthermore, we performed a novel statistical analysis with classification trees, which allowed us to depict a predictive core of nuclear receptor expression patterns characterizing subjects with Metabolic Syndrome. Random Forest Analysis identified NOR1 and PPARδ, which were both reduced in peripheral blood mononuclear cells and specifically in CD14(+) cells (mostly monocytes), as classifiers of Metabolic Syndrome, with high specificity and sensitivity. CONCLUSIONS: Our results point to the use of PPAR and NR4A mRNA levels in the overall peripheral blood mononuclear cells as biomarkers of Metabolic Syndrome and bona fide putative targets of pharmacological therapy.

Liver X receptors inhibit proliferation of human colorectal cancer cells and growth of intestinal tumors in mice.

BACKGROUND & AIMS: Liver X receptors (LXRs) are transcriptional regulators of cholesterol metabolism, controlling cholesterol flow into cells, catabolism, and efflux. Cholesterol controls cell proliferation; disruptions in cholesterol metabolism have been associated with the development of colon cancer. We investigated whether expression of activated LXR protects against intestinal tumorigenesis in mice. METHODS: We analyzed the development of colon cancer in mice that express a constitutive active form of LXRα only in the intestinal epithelium, under the control of villin promoter (iVP16LXRα). These mice were crossed with adenomatous polyposis coli (Apc)(min/+) mice, or given azoxymethane followed by dextran sodium sulfate, to assess intestinal tumor formation. We also assessed proliferation and apoptosis of a human colorectal cancer cell line (HT29) transfected with an adenoviral vector that expressed Ad VP16hLXRα, compared with cells expressing AdVP16 (control), and their ability to form xenograft tumors in mice. HT29 cells also were incubated with the LXR ligand GW3965. RESULTS: In human colorectal cancer cells, ligand-induced activation of LXR or transfection with Ad VP16hLXRα blocked the G1 phase, increased caspase-dependent apoptosis, and slowed growth of xenograft tumors in mice. iVP16LXRα mice formed fewer, smaller tumors than VP16 (control) mice after administration of azoxymethane and dextran sodium sulfate. APC(min/+)/iVP16LXRα mice also developed fewer, smaller intestinal tumors than APC(min/+)/iVP16 mice. Gene expression analysis indicated that activation of LXRα affected lipid metabolic networks and increased cholesterol efflux in the intestine. CONCLUSIONS: Expression of activated LXRα blocks proliferation of human colorectal cancer cells and slows the growth of xenograft tumors in mice. It also reduces intestinal tumor formation after administration of chemical carcinogens, and in Apc(min/+) mice. LXR agonists therefore might be developed as therapeutic treatments for colorectal cancer.

Neuron-derived orphan receptor 1 promotes proliferation of quiescent hepatocytes.

BACKGROUND & AIMS: Studies of the transcriptional networks that regulate nuclear receptor-mediated proliferation of quiescent hepatocytes could lead to new information about liver growth and hepatoprotective strategies. METHODS: We used quantitative real-time PCR to analyze expression of neuron-derived orphan receptor 1 (Nor-1) and its target genes during liver regeneration after hepatectomy in mice, and in hepatocellular carcinoma (HCC) samples from patients. We used adenoviral vectors to express Nor-1 in normal liver (Ad/CMV/V5-Nor-1), or reduce its level with small hairpin RNAs (Ad/BLOCK-iT/Nor-1(small hairpin RNA)) after partial hepatectomy. RESULTS: Levels of Nor-1 messenger RNA and protein, and transcription of Nor-1 target genes (Ccnd1 and Vcam-1), increased during the late priming and proliferative phases of liver regeneration after partial hepatectomy. Levels of NOR-1 messenger RNA and transcription of its target gene CCND1 and of the NOR-1 subfamily member NUR-77 also increased in human HCC samples compared with paired HCC-free tissue. Ad-Nor-1(small hairpin RNA) reduced the hepatocyte proliferation after hepatectomy. Overexpression of Nor-1 in normal livers of mice induced proliferation of quiescent hepatocytes independently of interleukin-6 and tumor necrosis factor-α signaling. In gene expression profile analysis, Nor-1 altered expression of genes involved in the cell cycle, proliferation, and tumorigenesis. CONCLUSIONS: In mice, the orphan nuclear receptor Nor-1 activates proliferation of quiescent hepatocytes and is required for hepatocyte proliferation after partial hepatectomy. Nor-1 and its gene targets are also up-regulated in human HCC samples. Nor-1 activates a transcriptional program that induces hepatocyte proliferation independently of inflammatory signaling pathways.

Selective activation of nuclear bile acid receptor FXR in the intestine protects mice against cholestasis.

BACKGROUND & AIMS: Cholestasis is a liver disorder characterized by impaired bile flow, reduction of bile acids (BAs) in the intestine, and retention of BAs in the liver. The farnesoid X receptor (FXR) is the transcriptional regulator of BA homeostasis. Activation of FXR by BAs reduces circulating BA levels in a feedback mechanism, repressing hepatic cholesterol 7α-hydroxylase (Cyp7a1), the rate-limiting enzyme for the conversion of cholesterol to BAs. This mechanism involves the hepatic nuclear receptor small heterodimer partner and the intestinal fibroblast growth factor (FGF) 19 and 15. We investigated the role of activation of intestine-specific FXR in reducing hepatic levels of BAs and protecting the liver from cholestasis in mice. METHODS: We generated transgenic mice that express a constitutively active FXR in the intestine. Using FXR gain- and loss-of-function models, we studied the roles of intestinal FXR in mice with intrahepatic and extrahepatic cholestasis. RESULTS: Selective activation of intestinal FXR induced FGF15 and repressed hepatic Cyp7a1, reducing the pool size of BAs and changing the BA pool composition. Activation of intestinal FXR protected mice from obstructive extrahepatic cholestasis after bile duct ligation or administration of α-naphthylisothiocyanate. In Mdr2(-/-) mice, transgenic expression of activated FXR in the intestine protected against liver damage, whereas absence of FXR promoted progression of liver disease. CONCLUSIONS: Activation of FXR transcription in the intestine protects the liver from cholestasis in mice by inducing FGF15 expression and reducing the hepatic pool of BA; this approach might be developed to reverse cholestasis in patients.

Familial mediterranean fever: a fascinating model of inherited autoinflammatory disorder.

BACKGROUND: Familial Mediterranean fever (FMF) is a rare inherited autosomal recessive autoinflammatory disorder characterized by recurrent and self-limited episodes of fever and painful serositis, lasting 1-3 days. FMF occurs almost exclusively among ethnic groups of the Mediterranean basin, although cases have also been found in Japan and Korean populations. Diagnosis is based on clinical features, response to colchicine and genetic analysis. Novel drugs are emerging, allowing better management of colchicine-resistant/colchicine-intolerant patients. This review aims to attract the attention of the readers on differential diagnosis and management of patients with FMF. METHODS: The current state-of-the-art on FMF is outlined, with respect to epidemiological, genetic, pathophysiological and therapeutic characteristics, based on critical analysis of solid scientific literature. RESULTS: FMF is more frequent than it was thought before. The phenotypic expression of M694V is more severe than that of V726A. Patients with M694V/M694V homozygosity are exposed to a higher risk of developing renal amyloidosis, arthritis, dermatologic and oral lesions, higher fever and more frequent painful attacks. Life-long therapy with colchicine (1·0-2·4 mg/day) is effective and safe to prevent recurrent attacks and renal amyloidosis and to reverse proteinuria. In nonresponder patients, alternative novel approaches include interleukin-1 receptor antagonist anakinra and the interleukin-1 decoy receptor rilonacept. CONCLUSIONS: The prognosis of FMF is normal if AA amyloidosis is prevented. Colchicine remains the first-line therapy to treat pain and prevent amyloidosis. A follow-up should include clinical evaluation, therapeutic adjustments, measurement of serum amyloid A and proteinuria.

Oxidized LDL receptor 1 gene polymorphism in patients with metabolic syndrome.

AIMS: The lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), encoded by the OLR1 gene, has been implicated in the pathogenesis of atherosclerosis. We therefore evaluated the genotyping of OLR1 gene in a sample of 55 patients with Metabolic Syndrome, a clinical condition characterized by a high cardiovascular risk. METHODS AND PATIENTS: The genotyping of the LOX-1 was performed by polymerase chain reaction (PCR) analysis of the IVS4-14 A>G OLR1 polymorphism embedded within the OLR1 Linkage Disequilibrium block. Patients were assessed for routine serum parameters, microalbuminuria, insulin resistance (HOMA) and oxidative stress (thiobarbituric acid reactive substances, TBARs and thioredoxin). RESULTS: The allele or genotype distribution of the OLR1 IVS4-14 A>G was not statistically different between MS and controls subjects. A positive association was found between IVS4-14 GG genotype, microalbuminuria and fasting glycaemia as well as a higher frequency of type 2 diabetes, elevated microalbuminuria, fasting serum glucose and HOMA index in the same subjects. Thioredoxin values were higher in patients with MS but did not differ in relation to OLR1 IVS4-14 A>G genotype. The TBARs/Cholesterol ratio was higher in MS both in IVS4-14 GG and in IVS4-14 AG. CONCLUSION: IVS4-14 GG genotype seems to be related to glucose metabolism disturbance, elevated insulin level and lipid peroxidation in patients with MS.

The intestinal nuclear receptor signature with epithelial localization patterns and expression modulation in tumors.

BACKGROUND & AIMS: The WNT-adenomatous polyposis coli system controls cell fate in the intestinal epithelium, where compartment-specific genes tightly regulate proliferation, migration, and differentiation. Nuclear receptors are transcription factors functioning as sensors of hormones and nutrients that are known to contribute to colon cancer progression. Here we mapped the messenger RNA (mRNA) abundance and the epithelial localization of the entire nuclear receptor family in mouse and human intestine. METHODS: We used complementary high-resolution in situ hybridization and systematic real-time quantitative polymerase chain reaction in samples of normal distal ileum and proximal colon mucosa and tumors obtained from mouse and human adenomatous polyposis coli-initiated tumor models (ie, Apc(Min/+) mice and familial adenomatous polyposis patients) and in cellular models of human colon cancer. RESULTS: We first defined for each receptor an expression pattern based on its transcript localization in the distal ileum and the proximal colon. Then, we compared the mRNA levels between normal intestinal epithelium and neoplastic intestinal tissue. After analyzing the correspondence between mouse and human tumor samples plus genetically modified human colon cancer cells, we used complementary graphic and statistical approaches to present a comprehensive overview with several classification trees for the nuclear hormone receptor intestinal transcriptome. CONCLUSIONS: We defined the intestinal nuclear hormone receptor map, which indicates that the localization pattern of a receptor in normal intestine predicts the modulation of its expression in tumors. Our results are useful to select those nuclear receptors that could be used eventually as early diagnostic markers or targeted for clinical intervention in intestinal polyposis and cancer.

13C-breath tests for clinical investigation of liver mitochondrial function.

BACKGROUND: Mitochondria play a major role in cell energetic metabolism; therefore, mitochondrial dysfunction inevitably participates in or even determines the onset and progression of chronic liver diseases. The assessment of mitochondrial function in vivo, by providing more insight into the pathogenesis of liver diseases, would be a helpful tool to study specific hepatic functions and to develop rational diagnostic, prognostic and therapeutic strategies. DESIGN: This review focuses on the utility of breath tests to assess mitochondrial function in humans and experimental animals. RESULTS: The introduction in the clinical setting of specific breath tests may allow elegantly and noninvasively overcoming the difficulties caused by previous complex techniques and might provide clinically relevant information, i.e the effects of drugs on mitochondria. Substrates meeting this requirement are alpha-keto-isocaproic acid and methionine that are both decarboxylated by mitochondria. Long-and medium-chain fatty acids that are metabolized through the Krebs cycle, and benzoic acid which undergoes glycine conjugation, may also reflect the function of mitochondria. CONCLUSIONS: Breath tests to assess in vivo mitochondrial function in humans represent a potentially useful diagnostic and prognostic tool in clinical investigation.

Coordinate regulation of gallbladder motor function in the gut-liver axis.

Gallstones are one of the most common digestive diseases with an estimated prevalence of 10%-15% in adults living in the western world, where cholesterol-enriched gallstones represent 75%-80% of all gallstones. In cholesterol gallstone disease, the gallbladder becomes the target organ of a complex metabolic disease. Indeed, a fine coordinated hepatobiliary and gastrointestinal function, including gallbladder motility in the fasting and postprandial state, is of crucial importance to prevent crystallization and precipitation of excess cholesterol in gallbladder bile. Also, gallbladder itself plays a physiopathological role in biliary lipid absorption. Here, we present a comprehensive view on the regulation of gallbladder motor function by focusing on recent discoveries in animal and human studies, and we discuss the role of the gallbladder in the pathogenesis of gallstone formation.

Liver function as assessed by breath tests in patients with hepatocellular carcinoma.

BACKGROUND: Little is known on hepatic function in patients with hepatocellular carcinoma (HCC). Metabolic changes were explored in HCC patients before/after nonsurgical therapy. MATERIALS AND METHODS: HCV-related Child-Pugh A cirrhotic patients with (n = 37) or without HCC (n = 14) and healthy controls (n = 23) were enrolled. Subjects underwent breath testing with (13)C-methacetin or (13)C-ketoisocaproate for exploring microsomal and mitochondrial function, respectively. HCC patients repeated the tests 1-2, 30, and 180 d after radiofrequency ablation (n = 27, RFA) or transarterial chemoembolization (n = 10, TACE). RESULTS: At baseline, cirrhotic patients showed decreased methacetin demethylation capacity compared with controls (8.1 +/- 2.1 versus 13.7 +/- 1.3% cum. dose exhaled at 60 min, M +/- CI, P < 0.001) and minor changes in ketoisocaproate decarboxylation. HCC patients had methacetin demethylation comparable to cirrhotic subjects, but a significantly lower ketoisocaproate decarboxylation (8.5 +/- 1.0 versus 11.6 +/- 1.9% cum. dose exhaled at 60 min, P < 0.001). Methacetin metabolism was significantly decreased following TACE (-28%, P < 0.05) but not RFA. Ketoisocaproate decarboxylation was unaffected by TACE but decreased after RFA (-27%, P < 0.05). A recovery was noticed with ketoisocaproate as a probe after 1 and 6 mo (P < 0.003). HCC recurrence was associated with early decrease of ketoisocaproate decarboxylation. CONCLUSIONS: Liver mitochondrial function is decreased in cirrhotic patients with HCC suggesting a possible tumor-induced suppressant effect. RFA but not TACE appears to spare residual (microsomal) liver mass, but induces such a transient stunning effect on mitochondrial function. Improved mitochondrial function after 1 and 6 mo from RFA may represent an additional parameter of treatment efficacy. Breath test assessing liver function may have potential applications in HCC management.

Oxidative stress-induced risk factors associated with the metabolic syndrome: a unifying hypothesis.

Although the biochemical steps linking insulin resistance with the metabolic syndrome have not been completely clarified, mounting experimental and clinical evidence indicate oxidative stress as an attractive candidate for a central pathogenic role since it potentially explains the appearance of all risk factors and supports the clinical manifestations. In fact, metabolic syndrome patients exhibit activation of biochemical pathways leading to increased delivery of reactive oxygen species, decreased antioxidant protection and increased lipid peroxidation. The described associations between increased abdominal fat storage, liver steatosis and systemic oxidative stress, the diminished concentration of nitric oxide derivatives and antioxidant vitamins and the endothelial oxidative damages observed in subjects with the metabolic syndrome definitively support oxidative stress as the common second-level event in a unifying pathogenic view. Moreover, it has been observed that oxidative stress regulates the expression of genes governing lipid and glucose metabolism through activation or inhibition of intracellular sensors. Diet constituents can modulate redox reactions and the oxidative stress extent, thus also acting on nuclear gene expression. As a consequence of the food-gene interaction, metabolic syndrome patients may express different disease features and extents according to the different pathways activated by oxidative stress-modulated effectors. This view could also explain family differences and interethnic variations in determining risk factor appearance. This review mechanistically focused on oxidative stress events leading to individual disease factor appearance in metabolic syndrome patients and their setting for a more helpful clinical approach.

Gastrointestinal symptoms and motility disorders in patients with systemic scleroderma.

BACKGROUND: Studies on gastrointestinal symptoms, dysfunctions, and neurological disorders in systemic scleroderma are lacking so far. METHODS: Thirty-eight scleroderma patients (34 limited, 4 diffuse), 60 healthy controls and 68 dyspeptic controls were scored for upper and lower gastrointestinal symptoms (dyspepsia, bowel habits), gastric and gallbladder emptying to liquid meal (functional ultrasonography) and small bowel transit (H2-breath test). Autonomic nerve function was assessed by cardiovascular tests. RESULTS: The score for dyspepsia (mainly gastric fullness) was greater in scleroderma patients than healthy controls, but lower than dyspeptic controls who had multiple symptoms, instead. Scleroderma patients with dyspepsia had a longer disease duration. Fasting antral area and postprandial antral dilatation were smaller in scleroderma patients than dyspeptic and healthy controls. Gastric emptying was delayed in both scleroderma patients (particularly in those with abnormal dyspeptic score) and dyspeptic controls, who also showed a larger residual area. Despite gallbladder fasting and postprandial volumes were comparable across the three groups, gallbladder refilling appeared delayed in dyspeptic controls and mainly dependent on delayed gastric emptying in scleroderma. Small intestinal transit was also delayed in 74% of scleroderma and 66% of dyspeptic controls. Bowel habits were similar among the three groups. Autonomic neuropathy was not associated with dyspepsia, gastric and gallbladder motility and small intestinal transit. CONCLUSION: In scleroderma patients dyspepsia (mainly gastric fullness), restricted distension of the gastric antrum and diffuse gastrointestinal dysmotility are frequent features. These defects are independent from the occurrence of autonomic neuropathy.