Helicobacter pylori Exposure in Nausea and Vomiting of Pregnancy Increases Risk of Preterm Delivery.

Background: Hyperemesis gravidarum (HG), a severe form of nausea and vomiting in pregnancy (NVP), is a leading indication for hospitalization in the first trimester. NVP and HG are associated with Helicobacter pylori (HP) infection in non-United States cohorts. How HP exposure and NVP interact to affect metabolic disturbance and pregnancy outcomes is not known. Materials and Methods: We designed a retrospective cohort study relating HP and NVP to serum electrolyte laboratory results, preterm delivery, and infant birth weight. Single academic institution discovery and independent multi-institutional validation cohorts included pregnant subjects with an HP test result. Associations of HP, NVP, and pregnancy outcomes were assessed with odds ratio calculations, Student's t-tests, and multivariate logistic regression. Results: Among subjects with positive HP test results, the prevalence of hyperemesis gravidarum (HG) was 0.025 (66 of 2671) and NVP was 0.27 (710 of 2671). Subjects with negative HP had prevalence of HG 0.015 (165 of 10,960) and NVP 0.22 (2392 of 10,960). History of HP exposure increased risk of NVP, including HG (odds ratio 1.3, 95% CI 1.1-1.4). Patients with HP exposure had lower serum potassium (mean difference 0.1 mEq/L) and bicarbonate (mean difference 0.3 mEq/L) during pregnancy than HP-negative patients (p < 0.01). Serum potassium was lowest in subjects with both NVP and HP exposure (mean 3.5 mEq/L [3.4-3.6], p < 0.0001). HP exposure alone carried increased risk for preterm delivery (OR 1.3 [1.1-1.4]). NVP alone increased risk of preterm delivery (OR 2.8 [2.5-3.1]) including second trimester delivery (OR 2.2 [1.7-2.8]). In multivariate analysis, HP exposure in the setting of NVP further increased risk of preterm delivery (adjusted OR 1.4 [1.0-1.9], p = 0.03). Conclusions: H. pylori exposure and diagnosis of NVP are individually associated with metabolic disturbances and adverse pregnancy outcomes such as preterm labor and delivery, and their combination further increases risk in US populations.

Structural disruption of Ntox15 nuclease effector domains by immunity proteins protects against type VI secretion system intoxication in Bacteroidales.

Bacteroidales use type VI secretion systems (T6SS) to competitively colonize and persist in the colon. We identify a horizontally transferred T6SS with Ntox15 family nuclease effector (Tde1) that mediates interbacterial antagonism among Bacteroidales, including several derived from a single human donor. Expression of cognate (Tdi1) or orphan immunity proteins in acquired interbacterial defense systems protects against Tde1-dependent attack. We find that immunity protein interaction induces a large effector conformational change in Tde nucleases, disrupting the active site and altering the DNA-binding site. Crystallographic snapshots of isolated Tde1, the Tde1/Tdi1 complex, and homologs from Phocaeicola vulgatus (Tde2/Tdi2) illustrate a conserved mechanism of immunity inserting into the central core of Tde, splitting the nuclease fold into two subdomains. The Tde/Tdi interface and immunity mechanism are distinct from all other polymorphic toxin-immunity interactions of known structure. Bacteroidales abundance has been linked to inflammatory bowel disease activity in prior studies, and we demonstrate that Tde and T6SS structural genes are each enriched in fecal metagenomes from ulcerative colitis subjects. Genetically mobile Tde1-encoding T6SS in Bacteroidales mediate competitive growth and may be involved in inflammatory bowel disease. Broad immunity is conferred by Tdi1 homologs through a fold-disrupting mechanism unique among polymorphic effector-immunity pairs of known structure. IMPORTANCE Bacteroidales are related to inflammatory bowel disease severity and progression. We identify type VI secretion system (T6SS) nuclease effectors (Tde) which are enriched in ulcerative colitis and horizontally transferred on mobile genetic elements. Tde-encoding T6SSs mediate interbacterial competition. Orphan and cognate immunity proteins (Tdi) prevent intoxication by multiple Tde through a new mechanism among polymorphic toxin systems. Tdi inserts into the effector central core, splitting Ntox15 into two subdomains and disrupting the active site. This mechanism may allow for evolutionary diversification of the Tde/Tdi interface as observed in colicin nuclease-immunity interactions, promoting broad neutralization of Tde by orphan Tdi. Tde-dependent T6SS interbacterial antagonism may contribute to Bacteroidales diversity in the context of ulcerative colitis.

Isolated MLH1 Loss by Immunohistochemistry Because of Benign Germline MLH1 Polymorphisms.

PURPOSE: Mismatch repair (MMR) immunohistochemistry (IHC) is frequently used to inform prognosis, select (immuno-)therapy, and identify patients for heritable cancer syndrome testing. However, false-negative and false-positive MMR IHC interpretations have been described. MATERIALS AND METHODS: Following identification of discordant MMR IHC and DNA-based microsatellite instability testing in a patient with colorectal carcinoma, we retrospectively reviewed institutional archives to identify patient samples with similar discrepancies. RESULTS: We report a patient with metastatic colorectal carcinoma who initially received immunotherapy on the basis of apparent isolated loss of MLH1 by IHC; notably, MLH1 promoter hypermethylation was negative. Subsequent evaluation of neoplastic tissue on a DNA-based targeted next-generation sequencing panel demonstrated microsatellite stability, low tumor mutational burden, and a benign MLH1 variant, MLH1 p.V384D, accompanied by loss of heterozygosity. The constellation of findings and repeat MLH1 IHC demonstrating retained expression using a different antibody-clone, supported reclassification of the neoplasm as MMR-proficient. Immunotherapy was discontinued, and cytotoxic chemotherapy was initiated. This index case of apparent discordance between MMR IHC and DNA-based microsatellite instability prompted a retrospective review of institutional archives to identify patient samples with similar discrepancies. Further evaluation of neoplasms harboring MLH1 p.V384D with loss of heterozygosity revealed systematic antibody-dependent interference. The review also identified a second IHC-interference candidate, MLH1 p.A441T. CONCLUSION: This study confirms that rare germline polymorphisms can result in incorrect IHC results, potentially affecting selection of optimal therapy and the decision to pursue germline testing. This case further highlights the need for expert molecular pathologic review and communication between clinical and molecular oncology teams.

Self-activating G protein α subunits engage seven-transmembrane regulator of G protein signaling (RGS) proteins and a Rho guanine nucleotide exchange factor effector in the amoeba Naegleria fowleri.

The free-living amoeba Naegleria fowleri is a causative agent of primary amoebic meningoencephalitis and is highly resistant to current therapies, resulting in mortality rates >97%. As many therapeutics target G protein-centered signal transduction pathways, further understanding the functional significance of G protein signaling within N. fowleri should aid future drug discovery against this pathogen. Here, we report that the N. fowleri genome encodes numerous transcribed G protein signaling components, including G protein-coupled receptors, heterotrimeric G protein subunits, regulator of G protein signaling (RGS) proteins, and candidate Gα effector proteins. We found N. fowleri Gα subunits have diverse nucleotide cycling kinetics; Nf Gα5 and Gα7 exhibit more rapid nucleotide exchange than GTP hydrolysis (i.e., "self-activating" behavior). A crystal structure of Nf Gα7 highlights the stability of its nucleotide-free state, consistent with its rapid nucleotide exchange. Variations in the phosphate binding loop also contribute to nucleotide cycling differences among Gα subunits. Similar to plant G protein signaling pathways, N. fowleri Gα subunits selectively engage members of a large seven-transmembrane RGS protein family, resulting in acceleration of GTP hydrolysis. We show Nf Gα2 and Gα3 directly interact with a candidate Gα effector protein, RGS-RhoGEF, similar to mammalian Gα12/13 signaling pathways. We demonstrate Nf Gα2 and Gα3 each engage RGS-RhoGEF through a canonical Gα/RGS domain interface, suggesting a shared evolutionary origin with G protein signaling in the enteric pathogen Entamoeba histolytica. These findings further illuminate the evolution of G protein signaling and identify potential targets of pharmacological manipulation in N. fowleri.

Delivery of therapeutic carbon monoxide by gas-entrapping materials.

Carbon monoxide (CO) has long been considered a toxic gas but is now a recognized bioactive gasotransmitter with potent immunomodulatory effects. Although inhaled CO is currently under investigation for use in patients with lung disease, this mode of administration can present clinical challenges. The capacity to deliver CO directly and safely to the gastrointestinal (GI) tract could transform the management of diseases affecting the GI mucosa such as inflammatory bowel disease or radiation injury. To address this unmet need, inspired by molecular gastronomy techniques, we have developed a family of gas-entrapping materials (GEMs) for delivery of CO to the GI tract. We show highly tunable and potent delivery of CO, achieving clinically relevant CO concentrations in vivo in rodent and swine models. To support the potential range of applications of foam GEMs, we evaluated the system in three distinct disease models. We show that a GEM containing CO dose-dependently reduced acetaminophen-induced hepatocellular injury, dampened colitis-associated inflammation and oxidative tissue injury, and mitigated radiation-induced gut epithelial damage in rodents. Collectively, foam GEMs have potential paradigm-shifting implications for the safe therapeutic use of CO across a range of indications.

Centrizonal hepatocyte dropout in allograft liver biopsies: a clinicopathological study.

AIMS: Centrizonal hepatocyte dropout has been described in diverse liver pathologies, including viral hepatitis, venous outflow obstruction, and allograft cellular rejection. However, its clinical significance remains uncertain. METHODS AND RESULTS: We designed a clinicopathological study of 206 allograft liver biopsies with centrizonal hepatocyte dropout. Centrizonal hepatocyte dropout was associated most frequently with cellular rejection (n = 62), asymptomatic/protocol biopsies (n = 56), immediate post-transplantation biopsies (n = 21), biliary obstruction (n = 14), and viral hepatitis (n = 13). The differential diagnosis is informed by timing post-transplantation, biliary imaging and laboratory test results. 'Cholestatic' and 'hepatocytic' laboratory patterns were associated with biliary obstruction and cellular rejection, respectively. A mixed pattern peaking after biopsy was observed in viral hepatitis cases. In the context of cellular rejection, dropout was not associated with the time interval to normalisation of serum alanine aminotransferase (ALT), but was associated with shorter transplant-free survival (hazard ratio 4, P = 0.01) than that of histological severity-matched controls. In time zero allograft biopsies, time to ALT normalisation was prolonged (median, 15 versus 11 days, P = 0.002) in allografts with centrizonal dropout, with no effect on retransplant-free survival. CONCLUSIONS: Centrizonal hepatocyte dropout has low clinicopathological diagnostic specificity. However, it correlates with adverse clinical outcomes in allograft cellular rejection and time zero biopsies.

Neutrophilic inflammation in gallbladder carcinoma correlates with patient survival: A case-control study.

Gallbladder carcinoma is an uncommon malignancy with an overall 5-year survival of less than 5%. Gallbladder carcinoma has been strongly linked with cholelithiasis and chronic inflammation. Case reports and series have described cholecystitis with acute (neutrophilic) inflammation in association with gallbladder carcinoma, although a clear relationship to patient outcome has not been established. Our series included 8 cases of gallbladder carcinoma with high tumor-associated neutrophils (>25 per high power field) that were associated with shorter patient survival (Cox regression coefficient 6.2, p = 0.004) than age- and stage-matched controls. High tumor-associated neutrophils were not associated with gallbladder rupture/perforation or increased bacterial load measured by 16S PCR. Neutrophilic inflammation with gallbladder carcinoma correlates to shorter survival, independent of patient age and stage of carcinoma. The findings suggest that the degree of neutrophilic inflammation may have prognostic significance in specimens from patients with gallbladder carcinoma after cholecystectomy. Further studies with larger case numbers are needed to confirm and generalize these findings.

Hepatocellular carcinoma in primary sclerosing cholangitis and primary biliary cholangitis: a clinical and pathological study in an uncommon but emerging setting.

Primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC) are biliary tract pathologies with increased risk of HCC, although HCC is more commonly associated with viral hepatitis and steatohepatitis. HCC risk stratification in PBC/PSC populations may help select patients for surveillance. We hypothesized that metabolic syndrome associated diagnoses and co-morbid nonalcoholic fatty liver disease (NAFLD) may be risk factors for HCC in patients with PBC and PSC. We undertook a multi-institutional case control study of PSC (19 cases, 38 controls) and PBC (39 cases and controls) patients with advanced fibrosis, matched for known HCC risk factors of age and sex, who had native liver explant or resection specimens. In the PSC population, HCC risk was significantly associated with multiple metabolic syndrome associated diagnoses (OR 13, p = 0.02), hyperlipidemia (OR 29, p = 0.03), and obesity (OR 6.8, p = 0.01). In the PBC cohort, only type 2 diabetes was a risk factor for HCC (OR 4.7, p = 0.03). In the PSC cohort, thick fibrous septae were associated with HCC risk (OR 3.4, p = 0.04). No other pathologic features of the nonneoplastic liver were significantly associated with HCC, including features of NAFLD such as macrovesicular steatosis, pericellular fibrosis, and steatohepatitis. Metabolic syndrome associated diagnoses, specifically type 2 diabetes among PBC patients, is associated with HCC risk in patients with biliary type cirrhosis. However, we found no evidence that HCC risk is related to co-morbid NAFLD, indicating a likely distinct mechanism of metabolic syndrome-associated carcinogenesis in these populations.

Quantification of Human Epidermal Growth Factor Receptor 2 by Immunopeptide Enrichment and Targeted Mass Spectrometry in Formalin-Fixed Paraffin-Embedded and Frozen Breast Cancer Tissues.

BACKGROUND: Conventional HER2-targeting therapies improve outcomes for patients with HER2-positive breast cancer (BC), defined as tumors showing HER2 protein overexpression by immunohistochemistry and/or ERBB2 gene amplification determined by in situ hybridization (ISH). Emerging HER2-targeting compounds show benefit in some patients with neither HER2 protein overexpression nor ERBB2 gene amplification, creating a need for new assays to select HER2-low tumors for treatment with these compounds. We evaluated the analytical performance of a targeted mass spectrometry-based assay for quantifying HER2 protein in formalin-fixed paraffin-embedded (FFPE) and frozen BC biopsies. METHODS: We used immunoaffinity-enrichment coupled to multiple reaction monitoring-mass spectrometry (immuno-MRM-MS) to quantify HER2 protein (as peptide GLQSLPTHDPSPLQR) in 96 frozen and 119 FFPE BC biopsies. We characterized linearity, lower limit of quantification (LLOQ), and intra- and inter-day variation of the assay in frozen and FFPE tissue matrices. We determined concordance between HER2 immuno-MRM-MS and predicate immunohistochemistry and ISH assays and examined the benefit of multiplexing the assay to include proteins expressed in tumor subcompartments (e.g., stroma, adipose, lymphocytes, epithelium) to account for tissue heterogeneity. RESULTS: HER2 immuno-MRM-MS assay linearity was ≥103, assay coefficient of variation was 7.8% (FFPE) and 5.9% (frozen) for spiked-in analyte, and 7.7% (FFPE) and 7.9% (frozen) for endogenous measurements. Immuno-MRM-MS-based HER2 measurements strongly correlated with predicate assay HER2 determinations, and concordance was improved by normalizing to glyceraldehyde-3-phosphate dehydrogenase. HER2 was quantified above the LLOQ in all tumors. CONCLUSIONS: Immuno-MRM-MS can be used to quantify HER2 in FFPE and frozen BC biopsies, even at low HER2 expression levels.

Elevated White Blood Cell Count Does Not Predict Clostridium difficile Nucleic Acid Testing Results.

BACKGROUND: An elevated white blood cell count (WBC; >15 000/µL) is an established prognostic marker in patients with Clostridium difficile infection (CDI). Small observational studies have suggested that a markedly elevated WBC should prompt consideration of CDI. However, there is limited evidence correlating WBC elevation with the results of C. difficile nucleic acid amplification testing (NAAT). METHODS: Retrospective review of laboratory testing, outcomes, and treatment of 16 568 consecutive patients presenting to 4 hospitals over 4 years with NAAT and WBC testing on the same day. RESULTS: No significant relationship between C. difficile NAAT results and concurrent WBC in the inpatient setting was observed. Although an elevated WBC did predict NAAT results in the outpatient and emergency department populations (P < .001), accuracy was poor, with receiver-operator areas under the curve of 0.59 and 0.56, respectively. An elevated WBC (>15 000/µL) in CDI was associated with a longer median hospital length of stay (15.5 vs 11.0 days; P < .01), consistent with leukocytosis as a prognostic marker in CDI. NAAT-positive inpatients with elevated WBC were more likely to be treated with metronidazole and/or vancomycin (relative ratio, 1.2; 95% confidence interval [CI], 1.1-1.3) and die in the hospital (relative ratio, 2.9; 95% CI, 2.0-4.3). CONCLUSIONS: Although WBC is an important prognostic indicator in patients with CDI, an isolated WBC elevation has low sensitivity and specificity as a predictor of fecal C. difficile NAAT positivity in the inpatient setting. A high or rising WBC in isolation is not a sufficient indication for CDI testing.

An interbacterial DNA deaminase toxin directly mutagenizes surviving target populations.

When bacterial cells come in contact, antagonism mediated by the delivery of toxins frequently ensues. The potential for such encounters to have long-term beneficial consequences in recipient cells has not been investigated. Here, we examined the effects of intoxication by DddA, a cytosine deaminase delivered via the type VI secretion system (T6SS) of Burkholderia cenocepacia. Despite its killing potential, we observed that several bacterial species resist DddA and instead accumulate mutations. These mutations can lead to the acquisition of antibiotic resistance, indicating that even in the absence of killing, interbacterial antagonism can have profound consequences on target populations. Investigation of additional toxins from the deaminase superfamily revealed that mutagenic activity is a common feature of these proteins, including a representative we show targets single-stranded DNA and displays a markedly divergent structure. Our findings suggest that a surprising consequence of antagonistic interactions between bacteria could be the promotion of adaptation via the action of directly mutagenic toxins.

Duodenal intraepithelial lymphocytosis in Helicobacter pylori gastritis: comparison before and after treatment.

Our aims were to assess performance of duodenal intraepithelial lymphocyte counting for diagnosis of Helicobacter pylori (H. pylori) gastritis, and effects of eradication therapy on intraepithelial lymphocytosis. Paired duodenal and gastric biopsies from subjects with a pathologic diagnosis of H. pylori gastritis were reviewed. Higher duodenal intraepithelial lymphocyte counts were observed in 40 subjects with H. pylori gastritis (26 ± 5 per villus) than 52 subjects negative for H. pylori (12 ± 2 per villus). After successful eradication therapy, duodenal lymphocytes were indistinguishable from H. pylori-negative subjects, whereas they remained elevated after failed eradication therapy. This study confirms previous reports of increased duodenal intraepithelial lymphocytes in patients with concurrent Helicobacter pylori gastritis. Intraepithelial lymphocyte counts of > 15 per villus or > 10 per 100 enterocytes were predictive of infection. Duodenal lymphocytosis decreases significantly after successful eradication therapy but remains elevated when treatment fails.

A bacterial cytidine deaminase toxin enables CRISPR-free mitochondrial base editing.

Bacterial toxins represent a vast reservoir of biochemical diversity that can be repurposed for biomedical applications. Such proteins include a group of predicted interbacterial toxins of the deaminase superfamily, members of which have found application in gene-editing techniques1,2. Because previously described cytidine deaminases operate on single-stranded nucleic acids3, their use in base editing requires the unwinding of double-stranded DNA (dsDNA)-for example by a CRISPR-Cas9 system. Base editing within mitochondrial DNA (mtDNA), however, has thus far been hindered by challenges associated with the delivery of guide RNA into the mitochondria4. As a consequence, manipulation of mtDNA to date has been limited to the targeted destruction of the mitochondrial genome by designer nucleases9,10.Here we describe an interbacterial toxin, which we name DddA, that catalyses the deamination of cytidines within dsDNA. We engineered split-DddA halves that are non-toxic and inactive until brought together on target DNA by adjacently bound programmable DNA-binding proteins. Fusions of the split-DddA halves, transcription activator-like effector array proteins, and a uracil glycosylase inhibitor resulted in RNA-free DddA-derived cytosine base editors (DdCBEs) that catalyse C•G-to-T•A conversions in human mtDNA with high target specificity and product purity. We used DdCBEs to model a disease-associated mtDNA mutation in human cells, resulting in changes in respiration rates and oxidative phosphorylation. CRISPR-free DdCBEs enable the precise manipulation of mtDNA, rather than the elimination of mtDNA copies that results from its cleavage by targeted nucleases, with broad implications for the study and potential treatment of mitochondrial disorders.

Serology Is More Sensitive Than Urea Breath Test or Stool Antigen for the Initial Diagnosis of Helicobacter pylori Gastritis When Compared With Histopathology.

OBJECTIVES: To assess the concordance and performance characteristics of Helicobacter pylori laboratory tests compared with histopathology and to propose algorithms for the diagnosis of H pylori that minimize diagnostic error. METHODS: H pylori diagnostics were reviewed from a 12-year period within a health system (2,560 cases). Analyses were performed to adjust diagnostic performance based on treatment and consensus histopathologic diagnoses among pathologists. Markers of access to care, including test cancellation frequency and turnaround time, were assessed. Costs and performance of candidate noninvasive testing algorithms were modeled as a function of disease prevalence. RESULTS: Serum H pylori IgG demonstrated a higher sensitivity (0.94) than urea breath and stool antigen tests (0.64 and 0.61, respectively). Evidence of an advantage in access to care for serology included a lower cancellation rate. Interobserver variability was higher (κ = 0.34) among pathologists for cases with a discordant laboratory test than concordant cases (κ = 0.56). A model testing algorithm utilizing serology for first-time diagnoses minimizes diagnostic error. CONCLUSIONS: Although H pylori serology has modestly lower specificity than other noninvasive tests, the superior sensitivity and negative predictive value in our population support its use as a noninvasive test to rule out H pylori infection. Reflexive testing with positive serology followed by either stool antigen or urea breath test may optimize diagnostic accuracy in low-prevalence populations.

Loss of nectin-3 expression as a marker of tumor aggressiveness in pancreatic neuroendocrine tumor.

Pancreatic neuroendocrine tumors (PanNETs) are rare, and prediction of aggressive characteristics, such as recurrence and metastasis and prognosis of PanNETs remain difficult. Nectins are cell adhesion molecules that regulate the formation of adherens and tight junctions. In this study, we investigated the clinicopathological significance of nectin-3 expression in patients with PanNETs. Immunohistochemical analysis of nectin-3 expression was performed on 78 cases of PanNET. Low nectin-3 expression in the membrane (positive ratio ≤25%) was observed in 62 cases (79.5%) and was significantly correlated with larger tumor size (>20 mm; P = 0.003), G2/G3 tumors (P = 0.025), higher Ki67 labeling index (≥3%; P = 0.009), lymphatic involvement (P = 0.047), advanced pT-factor (T2-T4; P = 0.003), lymph node metastasis (P = 0.006), advanced Union for International Cancer Control/American Joint Committee on Cancer-stage (Stage II-IV; P = 0.001), advanced ENETS stage (Stage IIa-IV; P = 0.001), nonfunctioning tumors (P = 0.002), and a shorter disease-free survival (P = 0.019). However, there was no significant correlation between nectin-3 expression in the membrane and/or cytoplasm and the clinicopathological parameters. The present results suggest that decreased nectin-3 expression in the membrane is associated with increased tumor aggressiveness of PanNETs. Clinically, immunohistochemical analysis of nectin-3 may help predict tumor aggressiveness for PanNETs.

Role of hepatocyte nuclear factor 4-alpha in gastrointestinal and liver diseases.

Hepatocyte nuclear factor 4-alpha (HNF4α) is a highly conserved member of nuclear receptor superfamily of ligand-dependent transcription factors that is expressed in liver and gastrointestinal organs (pancreas, stomach, and intestine). In liver, HNF4α is best known for its role as a master regulator of liver-specific gene expression and essential for adult and fetal liver function. Dysregulation of HNF4α expression has been associated with many human diseases such as ulcerative colitis, colon cancer, maturity-onset diabetes of the young, liver cirrhosis, and hepatocellular carcinoma. However, the precise role of HNF4α in the etiology of these human pathogenesis is not well understood. Limited information is known about the role of HNF4α isoforms in liver and gastrointestinal disease progression. There is, therefore, a critical need to know how disruption of the expression of these isoforms may impact on disease progression and phenotypes. In this review, we will update our current understanding on the role of HNF4α in human liver and gastrointestinal diseases. We further provide additional information on possible use of HNF4α as a target for potential therapeutic approaches.

Validation of a Congestive Hepatic Fibrosis Scoring System.

Congestive hepatopathy is a complication of right heart failure and chronically elevated right heart pressure. Histologic findings include sinusoidal dilatation, centrilobular hepatocellular plate atrophy, and fibrosis. We performed a validation study of a recently proposed scoring system (0 to 4 scale) for congestive hepatic fibrosis on 38 liver biopsies. Glutamine synthetase immunohistochemistry was also performed, and loss of centrizonal immunoreactivity correlated with increasing fibrosis score (P<0.01). Interobserver concordance for congestive hepatic fibrosis score based on Masson trichrome stain was initially fair (Fleiss κ=0.28, weighted concordance coefficient=0.60) and significantly improved (κ=0.40, weighted concordance coefficient=0.66) following a multiheaded microscope training session and inclusion of glutamine synthetase immunohistochemistry. Average congestive hepatic fibrosis score correlated with significantly higher right atrial pressure, severity of right atrial dilation, presence of right ventricular dilation, elevated serum alanine aminotransferase, platelet counts, prothrombin time, and model for end-stage liver disease score. In conclusion, the congestive hepatic fibrosis scoring system is reproducible among pathologists and correlates with clinical and laboratory markers of congestive hepatopathy.