Antonio De Vincentis

Breath-print analysis by e-nose for classifying and monitoring chronic liver disease: A proof-of-concept study

Since the liver plays a key metabolic role, volatile organic compounds in the exhaled breath might change with type and severity of chronic liver disease (CLD). In this study we analysed breath-prints (BPs) of 65 patients with liver cirrhosis (LC), 39 with non-cirrhotic CLD (NC-CLD) and 56 healthy controls by the e-nose. Distinctive BPs characterized LC, NC-CLD and healthy controls, and, among LC patients, the different Child-Pugh classes (sensitivity 86.2% and specificity 98.2% for CLD vs healthy controls, and 87.5% and 69.2% for LC vs NC-CLD). Moreover, the area under the BP profile, derived from radar-plot representation of BPs, showed an area under the ROC curve of 0.84 (95% CI 0.76–0.91) for CLD, of 0.76 (95% CI 0.66–0.85) for LC, and of 0.70 (95% CI 0.55–0.81) for decompensated LC. By applying the cut-off values of 862 and 812, LC and decompensated LC could be predicted with high accuracy (PPV 96.6% and 88.5%, respectively). These results are proof-of-concept that the e-nose could be a valid non-invasive instrument for characterizing CLD and monitoring hepatic function over time. The observed classificatory properties might be further improved by refining stage-specific breath-prints and considering the impact of comorbidities in a larger series of patients.

Lysosomal Acid Lipase Activity Is Reduced Both in Cryptogenic Cirrhosis and in Cirrhosis of Known Etiology.

Lysosomal acid lipase deficiency (LAL-d) is a rare autosomal recessive disease in which LAL activity is almost absent, with consequent massive microvesicular steatosis evolving to cirrhosis and liver failure. We aimed to determine LAL-activity, and to investigate the most common single nucleotide polymorphism (SNP) affecting the LIPA gene and responsible for 50–70% of LAL-d cases (rs116928232 c.894G>A), in patients with cryptogenic cirrhosis. Sixty-three patients with cryptogenic cirrhosis, 88 cirrhotics of known etiology, and 97 healthy subjects were enrolled. LAL-activity was determined in dried-blood-spot (DBS). The c.894G>A mutation was analyzed by pyrosequencing method in SNP mode. LAL-activity was severely reduced in patients with cryptogenic cirrhosis with respect to healthy subjects [0.62 (0.44–0.86) Vs 0.96 (0.75–1.25) nmol/spot/h, p<0.001)], but it was also reduced in known-etiology cirrhotics [0.54 (0.42–0.79) nmol/spot/h, p<0.001 Vs healthy subjects; p = 0.5 Vs cryptogenic cirrhotics]. Fourteen percent of cryptogenic cirrhotics and 20% of known-etiology cirrhotics showed a LAL-activity in the range of heterozygous carriers of LIPA gene mutations (0.15–0.40 nmol/spot/h). However, none of the subjects with reduced LAL-activity carried the c.894G>A SNP except for one patient with HCV cirrhosis. By multivariate analysis, LAL-activity was not associated with age, sex, liver enzymes, liver function or lipid parameters, while it was independently associated with white blood cell (β = 0.2; p<0.01) and platelet (β = 0.4; p<0.001) counts and with the condition of cirrhosis (β = -0.2; p = 0.04). Conclusion Liver cirrhosis is characterized by a severe acquired reduction of LAL-activity, the precise causes and consequences of which need to be further addressed. DBS-determined lysosomal enzyme activities seem to be affected by white blood cell and platelet counts, and the specificity of these tests can be reduced when applied to determined populations, such as cirrhotics.

Determinants of alanine aminotransferase levels in a large population from Southern Italy: Relationship between alanine aminotransferase and age

BACKGROUND Determinants of alanine aminotransferase levels have never been investigated in real-life settings. The relationship between alanine aminotransferase and age remains controversial. We evaluated epidemiological, anthropometric, and metabolic factors associated with alanine aminotransferase, focusing on the relationship between alanine aminotransferase and age. METHODS A 5-year retrospective analysis was performed on data recorded by 120 general practitioners from Naples (Italy), caring for 170,000 subjects. Exclusion criteria were age <18 years, diagnosis of chronic liver disease, positive markers for viral hepatitis, alcohol abuse, and alanine aminotransferase >100UI/L. RESULTS 44,232 subjects were enrolled (42.7% males, mean age 56±18 years). Alanine aminotransferase showed independent direct associations with body mass index, glycaemia, cholesterol, and triglycerides (p<0.001), and inverse associations with high-density lipoprotein cholesterol (p<0.001) and creatinine (p<0.01). The relationship between alanine aminotransferase and age was better expressed by polynomial regression (r=0.18, p<0.001), creating an inverted parabola. Mean alanine aminotransferase increased until the third decade in males and the fifth in females, with a subsequent progressive decrease in both genders. The inverse association between alanine aminotransferase and age in older subjects was independent from metabolic factors. CONCLUSIONS This real-life setting study, supports the concept that dysmetabolism is a strong determinant of liver injury. Based on our data, a reduction of the standard upper limit of normal alanine aminotransferase should be considered for older subjects.

Low Alanine Aminotransferase Levels in the Elderly Population: Frailty, Disability, Sarcopenia, and Reduced Survival

Background Although low alanine aminotransferase (ALT) levels have been associated with poor outcomes in the elderly population, the determinants subtending this association have been poorly explored. To gain insight into this topic, we analyzed data from a prospective population-based database (InCHIANTI study) in which frailty, disability, sarcopenia, and pyridoxine levels were systematically assessed. Methods Data are from 765 participants aged more than 65 years (mean age 75.3 years, women 61.8%), without chronic liver disease, malignancies, or alcohol abuse. Frailty was defined according to Fried criteria, sarcopenia through peripheral Quantitative-Computed-Tomography (lowest gender-specific tertile of the residuals of a linear regression of muscle mass from height and fat mass), and disability as self-reported need for help in at least one basic daily living activity. Associations of ALT with overall and cardiovascular mortality were assessed by Cox-models with time-dependent covariates. Results ALT activity was inversely associated with frailty, sarcopenia, disability, and pyridoxine deficiency; however, higher ALT was confirmed to be protective with respect of overall and cardiovascular mortality even in multiple-adjusted models including all these covariates (overall: hazard ratio [HR] 0.98 [0.96-1], p = .02; cardiovascular: 0.94 [0.9-0.98], p < .01). The association between ALT activity and mortality was nonlinear (J-shaped), and subjects in the lower quintiles of ALT levels showed a sharply increased overall and cardiovascular mortality. Conclusions These results suggest that reduced ALT levels in older individuals can be considered as a marker of frailty, disability, and sarcopenia, and as an independent predictor of adverse outcomes. The possible relationship between reduced ALT and impaired hepatic metabolic functions should be explored.

The PNPLA3 rs738409 C > G polymorphism is associated with the risk of progression to cirrhosis in NAFLD patients

Abstract Background and aims: The patatin-like phospholipase domain-containing 3 (PNPLA3) rs738409 C > G single nucleotide polymorphism (SNP) has been associated with steatosis and fibrosis in previous NAFLD populations in which cirrhotic patients were very poorly represented. Since not all NAFLD with fibrosis evolve to cirrhosis, we investigated the specific risk of cirrhosis conferred in NAFLD patients by carrying this SNP. Methods: Three groups were studied: patients with NASH-cirrhosis; patients with biopsy-proven non-cirrhotic NAFLD; healthy subjects undergoing medicine check-ups. Epidemiological, anthropometric, and clinical data were collected, and the SNP was analyzed by pyrosequencing. Results: Sixty-one patients with NASH-cirrhosis, 60 with non-cirrhotic NAFLD, and 125 healthy controls were included. Frequency of the PNPLA3 minor (G) allele was increased in patients with NASH-cirrhosis compared with non-cirrhotic NAFLD and controls (allele frequency: 0.598 versus 0.367 versus 0.2, respectively, p < 0.001), and different between the latter two groups (p < 0.001). Three-quarters (74%) of NASH cirrhotics carried at least one G allele, and almost half of them (46%) were GG homozygous. By multivariate analysis in the NAFLD population, each copy of the G allele was associated with an almost doubling of the risk of cirrhosis [OR 1.8 (1.02–3.2)], while being GG homozygous with a tripled risk compared with being CC homozygous [3.01 (1.03–10.8)]. Conclusions: In NAFLD patients, carriage of the PNPLA3G allele, and particularly of the GG genotype, is significantly associated with the risk of cirrhotic evolution. If confirmed in larger series, these results would suggest that most of NASH cases require the contribution of an altered PNPLA3 function to progress until cirrhosis.

Cardiotrophin-1 is not associated with carotid or coronary disease and is inversely associated with obesity in patients undergoing coronary angiography.

Introduction Cardiotrophin-1 (CT-1) is a member of the interleukin-6 superfamily with known hypertrophic and protective actions upon cardiac myocytes. Although its effects on myocardial tissue and its role in hypertensive heart disease are well documented, there are no studies on CT-1 blood levels in patients with coronary artery disease. In this study we aimed to verify the relationships of serum CT-1 with vascular disease and metabolic parameters in a population of patients undergoing coronary angiography due to clinical indications. Material and methods Serum levels of CT-1 were investigated in a cohort of 81 consecutive patients (median age 68 years (95% CI: 64–71), 59 males) undergoing coronary angiography and carotid Doppler ultrasound. Exclusion criteria were: acute coronary syndrome, already-established ischemic cardiopathy, chronic inflammatory diseases and presence or past history of cancer. Results Levels of CT-1 were inversely correlated with body mass index (BMI) and waist circumference (WC) (ρ = –0.261, p = 0.02; ρ = –0.224, p = 0.05, respectively). Moreover, obese patients showed significantly lower CT-1 concentrations than non-obese ones (1.18 (0.64–1.64) ng/ml vs. 1.56 (1.37–2.04) ng/ml, p = 0.013), and serum CT-1 was significantly reduced in patients with elevated compared to those with normal WC (1.43 (0.94–1.60) ng/ml vs. 1.64 (1.39–2.49) ng/ml, p = 0.047). Concentrations of CT-1 did not correlate either with the other parameters of metabolic syndrome or with markers of cardiovascular disease (carotid intima-media thickness, presence of carotid or coronary artery plaques). Conclusions Our results failed to demonstrate any association between CT-1 and carotid or coronary disease. The inverse association with BMI and WC fits with the latest experimental data on the role of CT-1 in dysmetabolic conditions and could help to further clarify the role of CT-1 in obesity and diabetes.

Haemorrhoidal disease in severe portal hypertension: a combined approach with transjugular intrahepatic portosystemic shunt (TIPS) and transanal haemorrhoidal dearterialization (THD).

Haemorrhoidal disease is a common finding in cirrhotic patients (40–44%) [1, 2], as haemorrhoidal plexus is a possible site of porto-systemic venous anastomosis. Portal hypertension (PH) can both exacerbate pre-existent essential haemorrhoidal varices (HV) and cause secondary vascular pathologies, called ano-rectal varices (ARV), particularly in conditions of severely increased hepatic vein pressure gradient (HVPG). Coexisting in up to 30% of cases, ARV are different from HV: as they are mainly due to portal pressure, they usually do not benefit from local therapies, requiring instead a corrective strategy for the underlying PH. Here we report a case of a 45-year-old man, known to have alcoholic Child-B liver cirrhosis, who was admitted to our clinic in March 2011 for severe anaemization (haemoglobin 5.8 g/dl) secondary to persistent rectorrhagia. On admission, he was haemodynamically stable and physical examination evidenced pale skin, tachycardia and tachypnoea, while laboratory results showed microcytic anaemia and severe thrombocytopenia (46 000/µl), and mild elevation of hepatic tests. On anamnestic deepening, he underwent an haemorrhoidal prolapsectomy 15 years earlier and a haemorrhoidal ligature 6 years earlier for persistent symptomatic bleeding haemorrhoids. Moreover, he had a 25-year history of severe alcohol consumption but he had been in a relatively good condition until August 2010, when he received the diagnosis of liver cirrhosis because of the first episode of hepatic decompensation (ascites and encephalopathy). Clinical examination evidenced third-degree haemorrhoids and a pancolonoscopy documented ARV excluding other colonic bleeding lesions, while upper endoscopy showed congestive gastropathy in absence of oesophageal varices. Doppler ultrasonography revealed an increased diameter of the main portal tract (13.5 mm), with preserved hepatopetal flow at reduced mean velocity (10 cm/s). An abdominal computed tomography scan with contrast medium detected a patent ectatic superior mesenteric vein with congestion of the rectal venous plexus and mild perihepatic ascites and splenomegaly (Figure 1). Repeated blood transfusions were prescribed to raise haemoglobin and propranolol in increasing dosage was started to reduce portal hypertension. However, severe rectorrhagia continued so we opted for reducing portal system congestion with a transjugular intrahepatic portosystemic shunt (TIPS). The initial portal venous pressure was 22 mm Hg and the estimated HVPG was 18 mm Hg. Despite reduction of the pressure gradient from 18 mm Hg to 4 mm Hg, rectorrhagia persisted, so surgical intervention for haemorrhoids was suggested. Transanal haemorrhoidal dearterialization (THD) was performed through a dedicated proctoscope, which incorporates a Doppler probe that allows one to identify and to ligate haemorrhoidal arteries and arteriovenous shunts. Transanal haemorrhoidal dearterialization is an innovative mini-invasive technique that has gradually gained in popularity among surgeons since the encouraging initial results reported by Morinaga et al. [3] in 1995. It consists of the Doppler-guided ligation of the terminal branches of the superior rectal artery, which solely contribute to the blood supply of the haemorrhoidal plexus [4, 5], thereby reducing the congestion [6]. The postoperative course was uneventful and the patient was discharged after 5 days. After a 15-month follow-up period, the patient is in good clinical condition, complying with his medical treatment, and the ano-rectal blood loss has not recurred. Figure 1 Coronal contrast-enhanced CT image showing ectatic superior mesenteric vein (white arrow) This case highlights how difficult it can be to diagnose and treat haemorrhoidal disease in cirrhotic patients with severe portal hypertension. The presence of persistent bleeding HV, relapsing after previous local therapies, and of endoscopic-evident ARV, suggested that the severely increased portal hypertension was the real cause of bleeding and of persistence of varices. So, we first reduced portal hypertension with both medical (β-blockers) and interventional (TIPS) techniques, probably achieving a reduction in ARV degree and, when we observed that rectorrhagia persisted, it was clear that blood loss was due to coexistent essential HV, this time. Thus, we treated them with a local mini-invasive technique (THD, previously described). Conventional surgical approaches are usually considered unsuitable in this context [7]: they can worsen portal hypertension after surgical removal of the venous porto-caval shunts, as well as having an increased risk of complications in cirrhotic patients. In conclusion, our experience sheds further light on the relevance of managing portal pressure in case of persistent bleeding and relapsing HV in presence of ARV: TIPS could be performed as the first approach in order to reduce the degree of portal hypertension and to plan, in case of ineffective resolution of ano-rectal bleeding, a mini-invasive surgical procedure with reduced operative risk.

Breath‐print analysis by e‐nose may refine risk stratification for adverse outcomes in cirrhotic patients

The spectrum of volatile organic compounds in the exhaled breath (breath‐print, BP) has been shown to characterize patients with cirrhosis and with worse hepatic function. However, the association of different BPs with clinically relevant outcomes has not been described yet. Hence, we aimed to evaluate the association between BPs, mortality and hospitalization in cirrhotic patients and to compare it with that of the “classical” prognostic indices (Child‐Pugh Classification [CPC] and MELD).

A case of Henoch-Schönlein purpura in the elderly: not just a 'second childhood'.

Henoch-Schönlein Purpura (HSP) is a small vessel-vasculitis that usually affects children and adolescents; its onset in adults is uncommon. We describe a case of HSP complicated with nephritis and extensive deep vein thrombosis in an 81-year-old Caucasian woman, successfully treated with oral corticosteroids. Even at the extremes of age, HSP should be considered in the differential diagnosis of leukocytoclastic vasculitis, with a particular attention to renal involvement, because of its potential morbidity and mortality in the elderly; in addition, ruling out an occult thrombotic event in course of HSP is mandatory, especially in the presence of additional thrombotic risk factors.

Combining Genetic Variants to Improve Risk Prediction for NAFLD and Its Progression to Cirrhosis: A Proof of Concept Study

Background & Aims Identifying NAFLD patients at risk of progression is crucial to orient medical care and resources. We aimed to verify if the effects determined by different single nucleotide polymorphisms (SNPs) could add up to multiply the risk of NAFLD and NASH-cirrhosis. Methods Three study populations, that is, patients diagnosed with NASH-cirrhosis or with noncirrhotic NAFLD and healthy controls, were enrolled. PNPLA3 rs738409, TM6SF2 rs58542926, KLF6 rs3750861, SOD2 rs4880, and LPIN1 rs13412852 were genotyped. Results One hundred and seven NASH-cirrhotics, 93 noncirrhotic NAFLD, and 90 controls were enrolled. At least one difference in allele frequency between groups was significant, or nearly significant, for the PNPLA3, TM6SF2, and KLF6 variants (p < 0.001, p < 0.05, and p = 0.06, resp.), and a risk score based on these SNPs was generated. No differences were observed for SOD2 and LPIN1 SNPs. When compared to a score of 0, a score of 1-2 quadrupled, and a score of 3-4 increased 20-fold the risk of noncirrhotic NAFLD; a score of 3-4 quadrupled the risk of NASH-cirrhosis. Conclusions The effects determined by disease-associated variants at different loci can add up to multiply the risk of NAFLD and NASH-cirrhosis. Combining different disease-associated variants may represent the way for genetics to keep strength in NAFLD diagnostics.