common variable immunodeficiency ( cvid ) and congenital x - linked agammaglobulinemia ( xla ) are the most common symptomatic primary antibody - deficiency syndrome ( pad ) . cvids are a heterogeneous group of hypogammaglobulinemia deficiencies of unknown aetiology , frequently diagnosed in adults . xla is inherited as an x - linked recessive trait and caused by b cell specific tyrosine - kinase ( btk ) gene mutations resulting in a bone marrow defective b cell differentiation . cvid patients are affected by recurrent infections in the airways and gastrointestinal tract , causing bronchitis , pneumonia , otitis , chronic sinusitis , and gastroenteritis . autoimmune disorders and granulomatous diseases the liver diseases in these conditions are poorly defined . before hcv screening test and viral inactivating steps in immunoglobulin ( ig ) production for replacement treatment , rna viruses were recognized as causes of acute and chronic liver diseases . later , a condition of nodular regenerative hyperplasia ( nrh ) has been described [ 711 ] . more recently , a condition known as idiopathic noncirrhotic portal hypertension ( incph ) whose morphological substrate is the presence of nrh was found to be associated with many immunological disorders [ 12 , 13 ] . in pad patients selected because of liver abnormalities , a high prevalence of cholestasis ( 65% ) and portal hypertension ( 50% ) liver biopsies revealed nonfibrosing architectural abnormalities consistent with nrh and anomalies of portal vessels . here , we describe the prevalence of spleno - portal axis abnormalities in an unselected cohort of patients of pad without known causes of liver diseases . an unselected cohort of 123 patients with a pad diagnosis ( 117 affected by cvid and 6 by xla ) based on the current diagnostic criteria of the european society for immune deficiencies / pan - american group for immune deficiencies ( esid / pagid )   were included in the study . all the 123 patients with pad have been regularly followed up in our referral centre for primary immune deficiencies . according to international guidelines for recipients of plasma derivatives ( polyvalent immunoglobulins ) every year all patients under replacement treatment underwent hiv rna , hcv rna , and hbsag detection . five cvid patients with concomitant hcv or hbv infection and one with histologically proven primary biliary cirrhosis were excluded . one patient with cvid was excluded for being affected by primary biliary cirrhosis , histologically proven . the mean follow - up time of the 117 patients ( 111 cvid and 6 xla ) analysed for the study was 18  14 years . a cumulative period of 1867 daily alcohol consumption was less than 30  g for men and 20  g for women . detailed information including personal data , pedigree , and age at diagnosis , immunological data , and clinical manifestations such as pneumonia , gastroenteritis , and autoimmune diseases were collected from medical files . all data were processed in a database and sent to cineca , an interuniversity computing centre responsible for processing and analysing the data . the following data were collected for each patient every 4 months following the italian pad guidelines ( http://www.aieop.org/ ) : platelet ( plt ) , red blood cell ( rbc ) , and white blood cell ( wbc ) counts ; chemistries : aspartate aminotransferase ( ast ) , alanine aminotransferase ( alt ) , alkaline phosphatases ( ap ) , gamma - glutamyl transferase ( ggt ) , total bilirubin , serum albumin , and prothrombin time . blood samples were collected once a year and tested by an enzyme immunoassay technique ( eia ) for detection of hepatitis b surface antigen ( hbsag ) ; hcv - rna was tested by using nested - pcr assays . chest computerized tomography ( ct ) scans were performed every four years according to the italian guidelines . upper gastrointestinal endoscopy with gastric and duodenal biopsies was performed every two years due to the increased risk of cvid - associated gastric cancer and lymphoproliferative diseases . abdominal ultrasonography ( us )   was performed every 24 years in all patients older than 18 years of age by the same investigator working in our centre . liver biopsy was performed in the five patients ( patients numbers 2 , 43 , 55 , 59 , and 72 in supplementary table 1 ; see supplementary material available online at http://dx.doi.org/10.1155/2014/672458 ) with esophageal varices or portosystemic collateral vessels to exclude liver cirrhosis . liver abnormalities : liver enzymes are aspartate aminotransferase ( ast ) > 40  ui / l , alanine aminotransferase ( alt ) > 40 ui / l , alkaline phosphatase ( ap ) > 129  ui / l , gamma glutamyl transferase ( ggt ) > 61 ui / l , and total bilirubin > 12  mol / l for at least for six consecutive months . splenomegaly occurs when spleen longitudinal axis is  12 cm in at least two consecutive exams . portal vein enlargement occurs when portal vein diameter is  > 13  mm in at least two consecutive ultrasound examinations ( measurements of the portal vein were obtained at its broadest point just distal to the union of the splenic and superior mesenteric veins ) . spleno - portal axis abnormalities are association between splenomegaly and portal vein enlargement . noncirrhotic portal hypertension is diagnosed according to the criteria defined by schouten et al . . bronchiectasis is shown as localized , irreversible dilation of part of the bronchial tree by ct scan . pneumonia 's symptoms and signs are consistent with an acute lower respiratory tract infection associated with new radiographic shadowing for which there was no other explanation ( e.g. , not pulmonary oedema or infarction ) . recurrent gastroenteritis has three or more episodes in a year of nausea , vomiting , diarrhoea , and abdominal cramps that could be accompanied by fever . lymphoid nodular hyperplasia ( lnh ) is the presence of lymphoid aggregates found in the small intestine identified by intestinal biopsy . autoimmune manifestations are diagnosed according to established criteria for rheumatoid arthritis , systemic lupus erythematosus , thyroiditis , sjgren syndrome , and autoimmune cytopenia . liver biopsy specimens were fixed in formalin , paraffin - embedded , and stained with hematoxylin eosin , masson 's trichrome , and picrosirius red and silver stain for reticulum . intrahepatic lymphocyte immunostaining was performed on paraffin sections using monoclonal antibodies directed against cd3 ( dako , glostrup , denmark ) . sinusoid endothelization was assessed with cd34 immunostaining with specific antibodies ( dako , glostrup , denmark ) . the kruskal - wallis one - way analysis of variance was used for comparison of data . one hundred and seventeen patients , 111 affected by cvid and 6 by xla , were included in the analysis . detailed data are provided in supplementary table 1 . a cumulative period of 1867 patients - year there is a general agreement that splenomegaly is a common feature in pad but its consequences are not well understood . here , we confirmed our previous data   showing a spleen enlargement in 71/117 patients ( 61% ) of our cohort . spleen diameter was highly correlated with portal vein diameter ( r2 = 0.5 ; p < 0.0001 ) ( figure 1 ) suggesting that an increased splenic venous flow secondary to splenomegaly could contribute to determining a condition of portal superflux . at ultrasound , 30 out of 117 patients ( 25.6% ) had signs of portal vein enlargement but only 1/6 of these had portal hypertension / incph , with portal systemic collaterals ( figure 2 ) . longitudinal assessment of abdominal ultrasounds demonstrated that portal vein enlargement and splenomegaly slightly increased in the observation period ( supplementary figure 1 ) . in the subgroup of patients without spleno - axis abnormalities , spleen and portal vein diameters remained within the normal ranges during the observational period . diameters of portal vein and liver were also significantly associated ( r2 = 0.34 , p = 0.0009 ) . spleno - portal axis abnormalities were invariably associated with a more severe pad phenotype ( table 2 ) with a higher prevalence of bronchiectasis ( p = 0.05 ) , gastroenteritis ( p = 0.0002 ) , lymphoid nodular hyperplasia ( p = 0.009 ) , and autoimmune manifestations ( p = 0.03 ) . the inverse correlation between platelet count and spleen diameter ( r2 = 0.22 ; p < 0.0001 ) suggests a mechanism of splenic sequestration as the main cause of thrombocytopenia in pad patients ( figure 1 ) . thirty patients ( 28 cvid , 2 xla ) had portal vein enlargement detected by ultrasounds , an indirect index of portal hypertension . four of these patients ( 3 cvid and 1 xla ) had esophageal varices ( 3 small and one large ) at upper gastrointestinal endoscopy and one patient had portal vein collaterals detected by ct scan . these five patients underwent liver biopsy , which excluded cirrhosis and thus they fulfilled the diagnostic criteria for incph . in the remaining 25 patients with portal vein enlargement without other clinical or radiological signs of portal hypertension the liver biopsy was not performed for ethical reasons . in patient with incph , histological findings consisted of liver sinusoidal congestive dilatation , presence of paraportal shunt vessels in the portal tract , and sinusoidal endothelization . parenchymal changes consisted of a micronodular transformation with nodules surrounded by an atrophic rim of hepatocytes in absence of fibrosis . representative aspects of architectural changes and sinusoidal inflammation observed in a pad patient ( patient number 2 ) are shown in figure 3 . in blood chemistry , all 5 patients ( 100% ) who fulfilled the criteria for incph had abnormalities in liver enzymes : 4 had signs of cholestasis and 1 showed an increase of transaminase levels ( supplementary table 1 ) . signs of cholestasis in the absence of incph were also detected in 1 patient with portal vein enlargement and splenomegaly , in 3 patients with isolated splenomegaly , and in 1 patient without spleno - portal abnormalities . two additional patients without spleno - portal abnormalities had a mild increase in transaminase levels of unclear origin . none of patients with incph had signs of liver failure defined as inr > 1.5 and albumin < 3.5 and none bled from varices . the only patient with large esophageal varices this procedure was effective since she had no bleeding episodes in the 3 years period after this treatment . however , two patients with incph ( one with cvid and one with xla ) died as a consequence of sepsis . we demonstrated that in comparison to pad patients without splenomegaly pad patients with splenomegaly have a decrease in the frequency of switched memory b cells ( 4.1  5.4% versus 7.9  6.9% ; p = 0.01 ) and an increased frequency of cd21 low b cells ( 20.1  18 versus 11.4  10 ; p = 0.02 ) confirming our previous data . abnormalities of t cell subsets frequencies were also observed in cd3+cd4 + cells ( 35.17  11.8% versus 42.7  11.15% ; p = 0.01 ) , in nave cd4 + cells ( 22.4  15.2% versus 44.18  19.1% ; p < 0.0001 ) , and in cd4 + memory t cells ( 81.2  13 versus 67.45  15.5 ; p = 0.0003 ) . more profound defects in b and t cell subsets were observed in patients with incph and/or portal vein enlargement . in fact , immunological abnormalities were more evident when we analysed pad patients according to their clinical phenotype : patients with spleno - portal axis abnormalities , patients with isolated splenomegaly , and patients without spleno - portal abnormalities ( figure 4 ) : lower frequencies of switched memory b cells ( 3.1  4.2% , versus 4.6  6.2 versus 7.6    6.6  p = 0.04 ) and cd4+cd45ra+cd62l+ nave t cells ( 14.8 + 12.3% versus 24.6  13.4 versus 44.3  19.5  p < 0.0001 ) and increased frequencies of cd4 + memory t cells ( 89.3  10% versus 78.5  15.7% versus 67  15.7 ; p = 0.00003 ) . severe abnormalities were also found in the frequencies of cd21 low cells that were increased ( 23.6  13.4% versus 20.1  18% versus 11.4  10 ; p = 0.02 ) and in the frequencies of cd4 + regulatory t cells that were decreased ( 1.6  2.3% versus 3.6  2% versus 3.5  1.6% ; p = 0.02 ) . seventeen patients had less than 15% of naive cd4 + t cells , a condition recently named late onset combined immune deficiency ( locid )   described in a subgroup of cvid patients at high risk of opportunistic infections . seven out of 17 locid patients had portal enlargement and 2 of these had incph . details of peripheral blood lymphocyte immunophenotyping with mean  sd , median , range , percentiles , and p values showing comparison between the 3 groups ( without splenic - axis abnormalities , with isolated splenomegaly and with spleno - portal abnormalities ) were shown in supplementary table 2 . this study shows that 25.6% patients with pad had signs of portal vein enlargement but that only 1/6 of these had portal hypertension / incph . the subgroup of patients with incph or spleno - portal abnormalities without signs of portal hypertension had severe clinical and immunological phenotypes with a severe defect of b cell terminal differentiation , t cell activation , and a deficiency in regulatory t cells . the hallmark of primary antibody deficiencies is a defect in the production of antibodies indispensable for the response against a wide variety of pathogens resulting in recurrent and severe infections . histological abnormalities have been found in primary and secondary lymphoid organs , including spleen , the largest secondary lymphoid organ , and in mucosa - associated lymphoid tissues . previous studies have demonstrated that the spleen is crucial in regulating immune homoeostasis , through its ability to link innate and adaptive immunity and to protect against infections . due to its unique structure and vascularisation , an increased spleen size is a common finding in pad patients but neither its causes nor its consequences are well understood . histological descriptions of spleen abnormalities after splenectomy showed granulomatous lesions , congestive red pulp , follicular hyperplasia , and atrophic germinal centres / white pulp . in pad , a reduction of a memory b cell subset developing in the spleen , called igm memory cells , is associated with the absence of secretory iga on intestinal epithelial cells . a defect in the intestinal immunity alters the mucosal permeability associated with an increased risk of bacterial infections as it happens also in other liver diseases . both reduction of memory b cells and very low iga levels are risk factors for various pad - associated conditions , including respiratory and gastrointestinal infections as well as splenomegaly [ 20 , 28 , 29 ] . a high prevalence up to 50% of gastrointestinal symptoms , mainly chronic diarrhoea due to intestinal infections caused by giardia lamblia , campylobacter jejuni , or salmonella spp.and malabsorption , has been described . here , we addressed the issue of the possible consequences of splenomegaly on the spleno - portal axis . we confirmed a high frequency of splenomegaly in more severe forms of pad , cvid , and xla . this is the first demonstration that portal abnormalities might occur also in xla , further strengthening our hypothesis on incph pathogenesis . splenomegaly might contribute to portal hypertension by increasing the portal flow ( portal superflux ) . in addition , the increased frequency of intestinal infections may increase the hepatic resistance by directly affecting the portal vein branches . differently from other studies that included only pad patients with abnormal liver function tests , we included in the study pad patients without known causes of liver diseases . when pad patients were selected because of functional liver abnormalities high frequencies of portal hypertension ( 50% ) and anicteric cholestasis ( 65% ) were observed . the histological finding consisted of nrh   associated with lymphocytic cd8 + cytotoxic t cells sinusoid lining disruption and portal vein endothelitis may imply a possible role of the intrasinusoidal infiltrating lymphocytes in the pathogenesis of the lesions . in our study at variance with previous reports [ 710 ] spleno - portal axis abnormalities were detected also in few pad patients without altered liver function tests . endoscopic band ligation to prevent the variceal bleeding was effective in the patient with large esophageal varices . this procedure should then be considered in pad patients in agreement with the guidelines for prevention of variceal bleeding . its success further supported the italian guidelines that recommend regularly performing an upper gastrointestinal endoscopy every two years in pad patients ( http://www.aieop.org/ ) . spleno - portal axis abnormalities might be the consequences of multiple inflammatory states including autoimmunity and infections . the immune disturbance is characterized by severe b cell defects , increased numbers of exhausted cd21low b cells [ 34 , 35 ] , and t cell activation point to an inflammatory or infectious state . furthermore , portal abnormalities were more frequent in the cvid group recently called locid , at higher risk of opportunistic infections , including gastrointestinal infections . in pad patients , infections , splenomegaly , increased blood venous flow , and inflammation , all , potentially contribute to liver damage . chronic infections and immunological abnormalities that are the hallmark of pad are also considered the main hypothesized pathophysiological mechanisms of incph [ 12 , 37 ] . in pad patients , despite the low prevalence , incph should be actively searched when signs of with portal hypertension are present . on the other hand , a diagnosis of pad
background and aim . 
 portal hypertension has been reported in association with acquired and primary immune deficiencies without a comprehensive description of associated spleno - portal axis abnormalities . 
 pathological mechanisms are poorly defined . 
 methods . 
 observational , single centre study with the aim of assessing the prevalence of spleno - portal axis abnormalities in an unselected cohort of 123 patients with primary antibody deficiencies and without known causes of liver diseases regularly followed up for a mean time of 18  14 years . 
 a cumulative period of 1867 patients - year was analysed . 
 clinical and immunological data , abdominal ultrasounds , ct scans , and endoscopy features were included in the analysis . results . 
 twenty - five percent of patients with primary antibody deficiencies had signs of portal vein enlargement but only 4% of them had portal hypertension , with portal systemic collaterals . 
 liver biopsies showed liver sinusoids congestive dilatation , endothelization , and micronodularity fulfilling the criteria for noncirrhotic portal hypertension . 
 patients with portal vein enlargement had severe clinical and immunological phenotypes . conclusions . in primary antibody deficient patients , infections , inflammations , splenomegaly , increased blood venous flow , and lymphocyte abnormalities contribute to establishment of liver damage possibly leading to noncirrhotic portal hypertension . 
 patients with primary antibody deficiency should be considered a good model to give insight into the pathological mechanisms underlying noncirrhotic portal hypertension .