Low Yield of Positive Cerebrospinal Fluid (CSF) Culture and Bacterial PCR Tests In Paediatric Patients With A Normal CSF White Cell Count.

The aim was to review paediatric patients who had a positive cerebrospinal fluid culture or bacterial PCR test, yet had a normal CSF white cell count for age. Patients were included if they had a CSF sample sent for culture (between 2005 and 2015) or bacterial PCR (2010-2015), however neurosurgical and neurology patients were excluded. Of the 2,482 patients reviewed, there were 101 patients with laboratory-confirmed bacterial meningitis included, of which 27 were positive by culture and PCR, 26 by culture alone and 48 were positive by PCR only. Eighteen (18%) of these patients had CSF white cell counts within normal range. Only one case with a normal CSF white cell count, where the lumbar puncture was done after six days of antibiotics, was deemed to be clinically significant. Bacterial PCR should not be routinely requested in patients with a normal CSF white cell count, unless their blood culture is positive or unless clinically indicated based on the assessment of a senior paediatrician.

Serological markers of extracellular matrix remodeling predict transplant-free survival in primary sclerosing cholangitis.

BACKGROUND: Primary sclerosing cholangitis is a progressive liver disease with a remarkably variable course. Biomarkers of disease activity or prognostic models predicting outcome at an individual level are currently not established. AIM: To evaluate the prognostic utility of four biomarkers of basement membrane and interstitial extracellular matrix remodeling in patients with primary sclerosing cholangitis. METHODS: Serum samples were available from 138 large-duct primary sclerosing cholangitis patients (of which 102 [74%] with IBD) recruited 2008-2012 and 52 ulcerative colitis patients (controls). The median follow-up time was 2.2 (range 0-4.3) years. Specific biomarkers of type III and V collagen formation (PRO-C3 and PRO-C5, respectively) and type III and IV collagen degradation (C3M and C4M, respectively) were assessed. The Enhanced Liver Fibrosis test, including procollagen type III N-terminal peptide, tissue inhibitor of metalloproteinase-1 and hyaluronic acid was assessed for comparison. RESULTS: All markers were elevated in primary sclerosing cholangitis compared to ulcerative colitis patients (P < 0.001). PRO-C3 showed the largest difference between the two groups with a threefold increase in primary sclerosing cholangitis compared to ulcerative colitis patients. Patients with high baseline serum levels of all markers, except C3M, had shorter survival compared to patients with low baseline serum levels (P < 0.001). Combining PRO-C3 and PRO-C5 the odds ratio for predicting transplant-free survival was 47 compared to the Enhanced Liver Fibrosis test's odds ratio of 11. CONCLUSIONS: Extracellular matrix remodeling is elevated in primary sclerosing cholangitis patients compared to ulcerative colitis patients. Furthermore, the interstitial matrix marker PRO-C3 was identified as a potent prognostic marker and an independent predictor of transplant-free survival in primary sclerosing cholangitis.

The good and the bad collagens of fibrosis - Their role in signaling and organ function.

Usually the dense extracellular structure in fibrotic tissues is described as extracellular matrix (ECM) or simply as collagen. However, fibrosis is not just fibrosis, which is already exemplified by the variant morphological characteristics of fibrosis due to viral versus cholestatic, autoimmune or toxic liver injury, with reticular, chicken wire and bridging fibrosis. Importantly, the overall composition of the ECM, especially the relative amounts of the many types of collagens, which represent the most abundant ECM molecules and which centrally modulate cellular functions and physiological processes, changes dramatically during fibrosis progression. We hypothesize that there are good and bad collagens in fibrosis and that a change of location alone may change the function from good to bad. Whereas basement membrane collagen type IV anchors epithelial and other cells in a polarized manner, the interstitial fibroblast collagens type I and III do not provide directional information. In addition, feedback loops from biologically active degradation products of some collagens are examples of the importance of having the right collagen at the right place and at the right time controlling cell function, proliferation, matrix production and fate. Examples are the interstitial collagen type VI and basement membrane collagen type XVIII. Their carboxyterminal propeptides serve as an adipose tissue hormone, endotrophin, and as a regulator of angiogenesis, endostatin, respectively. We provide an overview of the 28 known collagen types and propose that the molecular composition of the ECM in fibrosis needs careful attention to assess its impact on organ function and its potential to progress or reverse. Consequently, to adequately assess fibrosis and to design optimal antifibrotic therapies, we need to dissect the molecular entity of fibrosis for the molecular composition and spatial distribution of collagens and the associated ECM.

Fibrosis is not just fibrosis - basement membrane modelling and collagen metabolism differs between hepatitis B- and C-induced injury.

BACKGROUND: While morphological patterns differ, the molecular phenotype of liver fibrosis is considered a stereotypical response to chronic liver injury. However, with different cellular triggers and networks regulating fibrosis, the molecular responses of the injured liver may not be identical. AIM: To investigate whether differences in extracellular matrix (ECM) composition of the liver during fibrogenesis in two seemingly similar types of viral hepatitis could be reflected by differences in ECM turnover. METHODS: Utilising a cross-sectional design, we measured specific ECM protein fragments in plasma from 197 chronic hepatitis B (CHB) patients and 403 chronic hepatitis C (CHC) patients matched for inflammation grade and fibrosis stage. Markers of matrix metalloprotease degraded type I, III, IV and VI collagen (C1M, C3M, C4M, C6M) and type III and IV collagen formation (Pro-C3, P4NP7S). RESULTS: P4NP7S, C3M, C4M and C6M were significantly elevated in CHB compared to CHC. In contrast, Pro-C3 was significantly elevated in CHC compared to CHB. Pro-C3, C3M and C4M were increased in parallel with inflammation and fibrosis in both cohorts. C6M and P4NP7S were associated with inflammation and fibrosis only in CHC. Basement membrane collagen fragments P4NP7S and C4M were significantly higher in matched activity and fibrosis cohorts within CHB vs CHC. CONCLUSION: The main parameters to determine extracellular matrix biomarker levels are inflammation, fibrosis, and type of viral insult. Compared to CHC, CHB appears to induce a higher basement membrane turnover. This suggests that there are aetiology-dependent molecular signatures in liver fibrosis that could have pathogenic and diagnostic implications.

Circulating Elastin Fragments Are Not Affected by Hepatic, Renal and Hemodynamic Changes, But Reflect Survival in Cirrhosis with TIPS.

BACKGROUND AND AIMS: Progressive fibrosis increases hepatic resistance and causes portal hypertension with complications. During progressive fibrosis remodeling and deposition of collagens and elastin occur. Elastin remodeling is crucially involved in fibrosis progression in animal models and human data. This study investigated the association of circulating elastin with the clinical outcome in cirrhotic patients with severe portal hypertension receiving transjugular intrahepatic porto-systemic shunt (TIPS). METHODS: We analyzed portal and hepatic venous samples of 110 cirrhotic patients obtained at TIPS insertion and 2 weeks later. The circulating levels of elastin fragments (ELM) were determined using specific monoclonal ELISA. The relationship of ELM with clinical short-time follow-up and long-term outcome was investigated. RESULTS: Circulating levels of ELM showed a gradient across the liver before TIPS with higher levels in the hepatic vein. Interestingly, the circulating ELM levels remained unchanged after TIPS. The circulating levels of ELM in portal and hepatic veins correlated with platelet counts and inversely with serum sodium. Hepatic venous levels of ELM were higher in CHILD C compared to CHILD A and B and were associated with the presence of ascites. Patients with high levels of ELM in the hepatic veins before TIPS showed poorer survival. In multivariate analysis ELM levels in the hepatic veins and MELD were independent predictors of mortality in these patients. CONCLUSION: This study demonstrated that circulating levels of ELM are not associated with hemodynamic changes, but might reflect fibrosis remodeling and predict survival in patients with severe portal hypertension receiving TIPS independently of MELD.

Novel serological neo-epitope markers of extracellular matrix proteins for the detection of portal hypertension.

BACKGROUND: The hepatic venous pressure gradient (HVPG) is an invasive, but important diagnostic and prognostic marker in cirrhosis with portal hypertension (PHT). During cirrhosis, remodelling of fibrotic tissue by matrix metalloproteinases (MMPs) is a permanent process generating small fragments of degraded extracellular matrix (ECM) proteins known as neoepitopes, which are then released into the circulation. AIM: To investigate their potential as plasma markers for detection of PHT. METHODS: Ninety-four patients with alcoholic cirrhosis and 20 liver-healthy controls were included. Clinical and laboratory data of the patients were collected. All patients received HVPG measurement with blood sampling. In these samples, the following degradation or formation markers were measured: C1M (type I-collagen), C3M and PRO-C3 (type III collagen), C4M and P4NP 7S (type IV collagen), C5M (type V collagen), C6M (type VI collagen), BGM (biglycan), ELM (elastin), CRPM (CRP). RESULTS: All ECM markers except for CRPM correlated significantly with HVPG. Interestingly, C4M, C5M and ELM levels were significantly higher in patients with HVPG >10 mmHg. Multiple regression analysis identified PRO-C3, C6M and ELM as significant determinants, while the models A and B including PRO-C3, ELM, C6M and model for end-stage liver disease (MELD) provided better description of PHT (r = 0.75, P < 0.0001). The models provided odds ratios of >100 for having clinical significant PHT. CONCLUSIONS: These novel non-invasive extracellular matrix markers reflect the degree of liver dysfunction. The different degrees of portal hypertension correlated with these circulating neoepitopes. Using a single blood sample, these neoepitopes in combination with MELD detect the level of portal hypertension.

Self-reported antenatal adherence to medical treatment among pregnant women with Crohn's disease.

BACKGROUND: Adherence to medical treatment among women with Crohn's disease (CD) prior to and during pregnancy has never been reported. AIM: To examine both the predictors and prevalence rates of non-adherence to maintenance medical treatment among women with CD prior to and during pregnancy. METHODS: Among a population of 1.6 million inhabitants, we identified a total of 132 women with CD who had given birth during 2000-2005. Questionnaires were used to investigate predictors and extent of adherence. The validity of self-reported use of medication was assessed using data from the Danish Prescription Database. We used logistic regression to estimate prevalence odds ratios for non-adherence according to smoking status and other predictors. RESULTS: Eighty percent of the patients returned the questionnaire. A total of 58 (54%) women reported to have been on medical treatment, 50 of whom had fulfilled a prescription on relevant medication. Adherence to medical treatment was 72%. Fear of a negative effect on fertility/foetus was a reason for non-adherence by 18.8% prior to, and by 45.5% during, pregnancy. Among smokers, 30.8% were non-adherent compared with 11.5% among nonsmokers (prevalence odds ratio 3.41, 95% CI 0.8-14.7). CONCLUSIONS: Despite fear of a negative effect on fertility/foetus, adherence to medical treatment is high in women with CD. There is no substantial variation in adherence prior to and during pregnancy. Smoking prior to pregnancy is a predictor of non-adherence.

Azathioprine treatment during lactation.

BACKGROUND: Thiopurines are widely used to maintain remission in inflammatory bowel disease. Treatment during pregnancy is generally recommended to improve the chance of a normal birth outcome, but advice concerning breastfeeding is conflicting. Aim To estimate the exposure of breastfed infants to 6-mercaptopurine, as a metabolite of azathioprine, from maternal milk. METHODS: Eight lactating women with inflammatory bowel disease receiving maintenance therapy with azathioprine 75-200 mg daily were studied. Milk and plasma samples were obtained 30 and 60 min after drug administration and hourly for the following 5 h. RESULTS: The variation in the bioavailability of the drug was reflected in a wide range of peak plasma values of 6-mercaptopurine within the first 3 h. A similar curve, but with an hour's delay and at significantly lower concentrations varying from 2-50 microg/L, was seen in maternal milk. After 6 h an average of 10% of the peak values were measured. CONCLUSIONS: The major part of 6-mercaptopurine in breast milk is excreted within the first 4 h after drug intake. On the basis of maximum concentration measured, the infant ingests mercaptopurine of <0.008 mg/kg bodyweight/24 h. The findings confirm that breastfeeding during treatment with azathioprine seems safe and should be recommended, considering the extensive beneficial effects.

Pathology of the olfactory epithelium: smoking and ethanol exposure.

OBJECTIVE: To investigate the effects of tobacco smoke on the olfactory epithelium. Cigarette smoking has been associated with hyposmia; however, the pathophysiology is poorly understood. The sense of smell is mediated by olfactory sensory neurons (OSNs) exposed to the nasal airway, rendering them vulnerable to environmental injury and death. As a consequence, a baseline level of apoptotic OSN death has been demonstrated even in the absence of obvious disease. Dead OSNs are replaced by the mitosis and maturation of progenitors to maintain sufficient numbers of neurons into adult life. Disruption of this balance has been suggested as a common cause for clinical smell loss. This current study will evaluate the effects of tobacco smoke on the olfactory mucosa, with emphasis on changes in the degree of OSN apoptosis. STUDY DESIGN: A rat model was used to assess the olfactory epithelium after exposure to tobacco smoke. METHODS: Rats were exposed to tobacco smoke alone (for 12 weeks), smoke plus dietary ethanol (for the final 5 weeks), or to neither (control). Immunohistochemical analysis of the olfactory epithelium was performed using an antibody to the active form of caspase-3. Positive staining for this form of the caspase-3 enzyme indicates a cell undergoing apoptotic proteolysis. RESULTS: Control rats demonstrated a low baseline level of caspase-3 activity in the olfactory epithelium. In contrast, tobacco smoke exposure triggered a dramatic increase in the degree of OSN apoptosis that affected all stages of the neuronal lineage. CONCLUSIONS: These results support the following hypothesis: smell loss in smokers is triggered by increased OSN death, which eventually overwhelms the regenerative capacity of the epithelium.

Pneumolysin-induced complement depletion during experimental pneumococcal bacteremia.

To quantify complement depletion by pneumolysin during Streptococcus pneumoniae bacteremia, cirrhotic and control rats were infected intravenously with one of three isogenic mutant strains of S. pneumoniae expressing different forms of pneumolysin. Outcome measures included clearance of the organisms from the bloodstream, alterations in 50% serum hemolytic complement (CH(50)) activity and complement C3 levels during infection, and serum opsonic capacity at 18 h postinfection. Cirrhotic rats had significantly lower CH(50) and C3 levels than control rats, both before and after infection. However, initial complement levels did not predict bacterial load after 18 h of infection. Changes in CH(50) and C3 levels over the 18-h period correlated with numbers of H+C+ but not H+C- or PLY- organisms in the bloodstream at 18 h postinfection. The sera of cirrhotic rats infected with the H+C+ strain had significantly decreased levels of C3 and showed significantly lower opsonizing activity for S. pneumoniae than sera from H+C+-infected control rats. These studies suggest that under limiting concentrations of complement, the expression of pneumolysin by pneumococci has a significant, negative effect on serum complement levels and reduces serum opsonic activity.

Use of rat models to mimic alterations in iron homeostasis during human alcohol abuse and cirrhosis.

With alcoholism, there are marked disturbances in iron homeostasis that are linked to alterations in serum transferrin and ferritin concentrations. This study identifies rat models of alcohol abuse that closely mimic these disturbances. Male rats were placed in one of the following three protocols: (1) pair-feeding of liquid diets for 1-8 weeks; (2) agar-block feeding for 8 weeks; or (3) generation of cirrhosis with CCl(4). Serum samples were analyzed for ferritin, transferrin, and iron levels, and the transferrin iron saturation and ferritin/transferrin ratios were calculated. Liver iron concentrations were also determined. Serum transferrin levels were elevated in animals fed alcohol for 8 weeks in pair-feeding and agar-block feeding protocols, but reduced in rats with cirrhosis. Serum ferritin concentration was reduced in rats fed ethanol in the liquid diet, but increased in rats consuming ethanol in agar blocks, in rats pair-fed the liquid control diet, and in rats with cirrhosis. This finding was mirrored by liver nonheme iron concentrations in all experimental groups, but not in the corresponding control groups. Serum iron levels were significantly elevated only in rats fed the liquid control diet. There was a progressive decrease in transferrin iron saturation and ferritin/transferrin ratios for animals fed ethanol in the liquid diet, but not when ethanol was ingested from agar blocks. The development of cirrhosis resulted in elevated liver iron concentrations and doubled ferritin/transferrin ratios. It is concluded that these models may be used to study disturbances in iron homeostasis that occur during alcohol abuse and the (subsequent) development of liver disease.

Host-parasite relationship in Pneumocystis carinii infection: activation of the plasmalemmal vesicular system in type I alveolar epithelial cells.

Ultrastructural studies of the attachment zone between Pneumocystis carinii (Pc) and type I alveolar epithelial cells showed a new aspect of the host-parasite relationship, i.e. an activation of the plasmalemmal vesicular system in the alveolar cells associated with Pc trophozoites in close apposition. This phenomenon may be involved in the nutrition of the trophozoite.

Blood group ABO and Lewis antigen expression during neoplastic progression of human urothelium. Immunohistochemical study of type 1 chain structures.

The deletion of blood group ABO antigen expression in bladder carcinoma has attracted attention because of its potential as a prognostic parameter. Based on recently produced monoclonal antibodies against blood group antigens, it has become possible to elucidate the carcinoma-associated modulation of these antigens at a molecular level. In this study we have used a panel of monoclonal antibodies (H, Lea, Leb, A, ALeb) that are specific to type 1 chain structures. By the use of an immunohistochemical method, the histologic and cytologic location of these antigens in the urothelium was studied in 25 biopsies from transitional cell carcinomas and compared to 21 previously examined normal biopsies. Urothelial blood group reactivity was compared to Lewis and secretor status. The authors found a series of events associated with neoplastic progression of noninvasive urothelium: a disruption of the orderly stratification of blood group antigens in different cell layers; cytostructural relocation of cytoplasmic antigens to the cell surface; loss of correlation between urothelial blood group antigens and secretor status; and gradual deletion of antigens. In the invasive tissue these events were followed by a total deletion of A and H isoantigens and uniform expression of Lewis b and sialyted Lewis a antigen. These findings indicate that there is a complex modulation of blood group antigen biosynthesis associated with the neoplastic progression of the human urothelium.