Resistance mutations to lamivudine and/or ETV was detected only in 3 and 2 patients this website in the TDF and TDF+ETV groups, respectively, at 48 weeks. None developed additional resistance mutations. None in the TDF group required protocol-defined switch over of treatment. Both treatments were

well tolerated, and safety and adverse event profiles were similar in the two groups. Conclusions: TDF monotherapy showed similarly high antiviral efficacy and safety as TDF and ETV combination therapy during 48 weeks of treatment in patients with ETV-resistant HBV. None developed additional resistance mutations. KEY WORDS: Lami-vudine, Monotherapy, Resistance, Virologic response Disclosures: Young-Suk Lim – Advisory Committees or Review

Panels: Gilead Science, Bayer; Grant/Research Support: Gilead Science, Novartis, Bayer; Speaking and Teaching: BMS Kwan Soo Byun – Advisory Committees or Review Panels: Gilead; Grant/ Research Support: Gilead, BMS, Taiho, Jassen; Speaking and Teaching: BMS Han Chu Lee – Grant/Research Support: Medigen Biotechnology Co., Novartis, Roche, Bayer HealthCare, Bristol-Myers Squibb, INC research, Boehringer Ingel-heim, Taiho Pharmaceutical NVP-BGJ398 research buy Co., Yuhan Co. The following people have nothing to disclose: Geum-Youn Gwak, Byung Chul Yoo, So Young Kwon, Yoon Jun Kim, Jihyun An, Yung Sang Lee Background: Antiviral therapy may reduce HCC risk but it is unclear what the residual risk would be in treated patients. Our aim is to characterize PIK-5 HCC incidence in treated and untreated patients by cirrhosis status, age (< 45 or ≥45), and gender. Methods: In this retrospective cohort study, 3933 consecutive CHB patients were identified at 3 US centers from 1991-2014. Patients were included if they had at least one year of follow-up and treatment-naïve. Exclusion criteria included HCC at initial presentation and the development of HCC within the first year of follow-up. Diagnosis was based on AASLD criteria for HCC and histology or clinical or imaging data for cirrhosis. Annual

incidence was calculated in cases per 1000 person years. Results: We included a total of 3220 patients with 102 incident HCC cases over a median time of follow-up of 4.1 (1-17) years. In multivariate analysis, antiviral therapy was an independent predictor for reduced HCC risk (HR 0.43, 95% CI 0.23-0.79) following adjustment for age, gender, cirrhosis, HBeAg, ALT, and HBV DNA. In cirrhotic men, regardless of age, the treated group had a lower incidence of HCC (Figure 1). For the non-cirrhotic cohort, the effects of antivirals, while beneficial were modest with the exception of men ≥45 years of age. HCC incidence in treated non-cirrhotic patients ranged from 0 to 1.2 cases per 1000 person years among the various age and gender groups compared to 0 to 6.4 per 1000 person years if untreated. HCC incidence in cirrhotic patients still ranged 16.

5, 6 In the present study, adiponectin levels were not significantly elevated in those with advanced-stage NASH fibrosis/cirrhosis when compared to those with early disease. One might have expected the levels to be lower in patients with advanced NASH who were more insulin resistant and obese

than those with early disease, but it is established that adiponectin MAPK inhibitor levels in cirrhosis do not correlate with insulin resistance, dyslipidemia, or obesity.17, 18 The unaltered levels of adiponectin in late compared to early disease is in part a deliberate consequence of our strict selection criteria, wherein we excluded (1) all patients with markers of liver synthetic dysfunction such as abnormal prothrombin time, albumin, or bilirubin, and (2) those with Child’s B and C cirrhosis. Thus, we were able to exclude elevations due to these confounders

known to be associated with increased adiponectin, and further strengthen our hypothesis.17, 18 Adiponectin levels are also lower in patients with nonalcoholic fatty liver disease (NAFLD) compared to other liver diseases29 and levels decline further with increasing necroinflammation and fibrosis. Thus, the finding of similar adiponectin levels for our two groups is in keeping with a Natural Product Library manufacturer relative elevation of adiponectin, similar to that seen in other forms of cirrhosis. Taken together, these findings suggest that physiological regulation of adiponectin Carbachol is dramatically altered in patients with advanced-stage liver disease compared to the situation in healthy volunteers, diabetes, or early liver disease.16, 30 A number of mechanisms have been hypothesized to explain the relative elevation in adiponectin with progressive fibrosis, including an imbalance between adiponectin production and hepatic extraction,18, 31 a protective antiinflammatory mechanism in the chronic inflammatory

state of cirrhosis,18 and an increase in true hepatocyte or hepatic stellate cell adiponectin production.17, 32 Because the highest levels of adiponectin are seen in patients with advanced cholestatic liver disease, reduced biliary excretion of adiponectin may also be important.15, 17, 33 This theory is supported by bile duct ligation studies in mice where dramatic increases in serum adiponectin were seen over time, and the detection of adiponectin in the bile of human subjects with severe cholestasis.15 None of the patients included in this study were severely catabolic or clinically had cholestasis (elevations in bilirubin), which raised the intriguing question as to why adiponectin would be elevated in our cohort and whether there could be a link between hepatocyte dysfunction and adiponectin production by adipose tissues.

5, 6 In the present study, adiponectin levels were not significantly elevated in those with advanced-stage NASH fibrosis/cirrhosis when compared to those with early disease. One might have expected the levels to be lower in patients with advanced NASH who were more insulin resistant and obese

than those with early disease, but it is established that adiponectin Selleck Opaganib levels in cirrhosis do not correlate with insulin resistance, dyslipidemia, or obesity.17, 18 The unaltered levels of adiponectin in late compared to early disease is in part a deliberate consequence of our strict selection criteria, wherein we excluded (1) all patients with markers of liver synthetic dysfunction such as abnormal prothrombin time, albumin, or bilirubin, and (2) those with Child’s B and C cirrhosis. Thus, we were able to exclude elevations due to these confounders

known to be associated with increased adiponectin, and further strengthen our hypothesis.17, 18 Adiponectin levels are also lower in patients with nonalcoholic fatty liver disease (NAFLD) compared to other liver diseases29 and levels decline further with increasing necroinflammation and fibrosis. Thus, the finding of similar adiponectin levels for our two groups is in keeping with a www.selleckchem.com/screening/tyrosine-kinase-inhibitor-library.html relative elevation of adiponectin, similar to that seen in other forms of cirrhosis. Taken together, these findings suggest that physiological regulation of adiponectin selleck is dramatically altered in patients with advanced-stage liver disease compared to the situation in healthy volunteers, diabetes, or early liver disease.16, 30 A number of mechanisms have been hypothesized to explain the relative elevation in adiponectin with progressive fibrosis, including an imbalance between adiponectin production and hepatic extraction,18, 31 a protective antiinflammatory mechanism in the chronic inflammatory

state of cirrhosis,18 and an increase in true hepatocyte or hepatic stellate cell adiponectin production.17, 32 Because the highest levels of adiponectin are seen in patients with advanced cholestatic liver disease, reduced biliary excretion of adiponectin may also be important.15, 17, 33 This theory is supported by bile duct ligation studies in mice where dramatic increases in serum adiponectin were seen over time, and the detection of adiponectin in the bile of human subjects with severe cholestasis.15 None of the patients included in this study were severely catabolic or clinically had cholestasis (elevations in bilirubin), which raised the intriguing question as to why adiponectin would be elevated in our cohort and whether there could be a link between hepatocyte dysfunction and adiponectin production by adipose tissues.

Both treatment strategies were well tolerated. Disclosures: Qin Ning – Advisory Committees or Review Panels: ROCHE, NOVARTIS, BMS, MSD, GSK; Consulting: ROCHE, NOVARTIS, BMS, MSD, GSK; Grant/Research Support: ROCHE, NOVARTIS, BMS; Speaking and Teaching: ROCHE, NOVAR-TIS, BMS, MSD, GSK The following people have nothing to disclose: Wei Guo, Di Wu, Peng Wang, Jing-Feng Chen, Yan-hong Xu, Weiming Yan, Ke Ma, Meifang Han, Jianxin Song, Junying Qi, Jiaquan Huang, Yuancheng Huang, Xiping Zhao, Dong Xu Background/Aims: DNA Damage inhibitor The antiviral effect of monotherapy with tenofovir disoproxil fumarate (TDF) is controversial for patients with hepatitis B virus (HBV) resistant

to nucloes(t)ide analogues. Methods: Patients who were treated with TDF monotherapy for HBV with

documented genotypic resistance mutations were included. Results: At baseline, 325 (81%) patients had mono-resistance to lamivudine, while others had multi-drug resistance to lamivudine and adefovir (n=43, 10%) or lamivudine and ente-cavir (n=32, 8%). Most (287, 72%) were being treated with combination therapies including lamivudine, adefovir, and entecavir. Their mean HBV DNA level was 2.5 +/− 2.0 log10 IU/mL. During 1 year of TDF monotherapy, 378 (95%) patients cumulatively achieved virological response (VR, HBV DNA <60 IU/mL) by modified intention-to-treat analysis. The rates of VR were not different between patients with mono-resistance to lamivudine and those with www.selleckchem.com/products/GDC-0449.html multi-drug resistance (95.2% vs. 92.0%, P =0.27). Sixteen patients who did not achieve VR also showed a significant reduction in their mean HBV DNA levels from baseline (-3.42 log10 IU/mL, P <0.01). Five patients experienced viral breakthrough, and all were related with low adherence to medication. The rate of HBV DNA undetectabil-ity

was not influenced by the degree of previous resistance mutations (P >0.05). Four Nintedanib (BIBF 1120) patients discontinued TDF because of gastrointestinal symptoms (n=3) or for pregnancy (n=1). Otherwise, no significant clinical or laboratory adverse event was reported. Conclusions: In this cohort study, most patients infected with HBV that has genotypic resistance mutations to lamivudine, adefovir, or entecavir rapidly achieved VR within 48 weeks of TDF monotherapy, regardless of the nature of previous resistance mutations. No patient developed additional resistance mutations. Disclosures: Young-Suk Lim – Advisory Committees or Review Panels: Bayer Healthcare, Gilead Sciences; Grant/Research Support: Bayer Healthcare, BMS, Gilead Sciences, Novartis Han Chu Lee – Grant/Research Support: Medigen Biotechnology Co., Novartis, Roche, Bayer HealthCare, Bristol-Myers Squibb, INC research, Boehringer Ingelheim, Taiho Pharmaceutical Co., Yuhan Co.

9% lower social class and

5% others. The average family salary was 500 dollars/monthly. 95.2% were literate. 16% had internet access. 9% read newspapers daily, 27% read sometimes, 26% read rarely and 38% never read newspapers. The informed age of onset of sexual activity was 1 6.4±3,4 years. Regarding number of partners/year, 57.7% reported one, 25.7% among 2–5 and 5.8% more than 5 partners/year. 46% reported regular use of condom, 27% irregular use and 27% never use it. Previous sexually transmitted diseases occurred in 19%. 76.1% reported regular share of Selleck Gefitinib at least one personal item. 53% and 3.5% share cuticle nippers and toothbrush, respectively, 20% share razors blade at home and 8% in barbershops. Overall, 1 6% used unsafe glass syringes/needles in the last 20 years. 3090 (52.7%) reported knowing about hepatitis. Hepatitis B was the most commonly reported (97%), followed by hepatitis A (87%) and C (75%). Only 27% knew the risk factors for viral hepatitis. The sources of information were TV (72%), newspapers, magazines or books (60%), schools (58%), public primary care (55%) or government advices (42%), person-to-person (35%), family experience (20%) and internet (16%). CONCLUSIONS: The knowledge about viral PD98059 research buy hepatitis and its risk factors in Minas Gerais

(Brazil) is low. The prevalence of sharing personal items and unsafe sexual activity is high. These data reinforce the need of establishing effective government actions aiming the prevention of viral hepatitis in Brazil and may alert other countries in similar conditions. Disclosures: The following people have nothing to disclose: Rosangela Teixeira, Filipe M. Araújo, Emilio also Suyama, Maria Aparecida M. Pereira, Geraldo Scarabelli, Soraia Z. Morais Introduction: In 2012, the American Board of Internal Medicine (ABIM) in collaboration with the American Association of the Study of Liver Diseases (AASLD) approved a competency-based Transplant Hepatology (TH) training pilot program. This program allows completion of both Gastroenterology (GI) and TH training in

three years. The institution of this pilot is a milestone towards a competency-based education model for training programs. The aim of this study was to identify the perceptions and beliefs of GI/Hepatology Division and Fellowship Program Directors on the combined GI/TH training pilot and competency-based education in GI fellowship. Methods: A 21 item survey was created to assess perceptions/beliefs about the 3-year combined GI/TH training pilot and the level of competency of graduates from the program. All current GI/TH Division and Fellowship Program Directors from AGCME-accredited programs were invited, via email to the Director, to anonymously complete the online survey (SurveyMonkeyTM). Results: A total of 1 1 6 participants completed the survey with a ~38% response rate.

9% lower social class and

5% others. The average family salary was 500 dollars/monthly. 95.2% were literate. 16% had internet access. 9% read newspapers daily, 27% read sometimes, 26% read rarely and 38% never read newspapers. The informed age of onset of sexual activity was 1 6.4±3,4 years. Regarding number of partners/year, 57.7% reported one, 25.7% among 2–5 and 5.8% more than 5 partners/year. 46% reported regular use of condom, 27% irregular use and 27% never use it. Previous sexually transmitted diseases occurred in 19%. 76.1% reported regular share of Selleckchem GSK458 at least one personal item. 53% and 3.5% share cuticle nippers and toothbrush, respectively, 20% share razors blade at home and 8% in barbershops. Overall, 1 6% used unsafe glass syringes/needles in the last 20 years. 3090 (52.7%) reported knowing about hepatitis. Hepatitis B was the most commonly reported (97%), followed by hepatitis A (87%) and C (75%). Only 27% knew the risk factors for viral hepatitis. The sources of information were TV (72%), newspapers, magazines or books (60%), schools (58%), public primary care (55%) or government advices (42%), person-to-person (35%), family experience (20%) and internet (16%). CONCLUSIONS: The knowledge about viral buy Smoothened Agonist hepatitis and its risk factors in Minas Gerais

(Brazil) is low. The prevalence of sharing personal items and unsafe sexual activity is high. These data reinforce the need of establishing effective government actions aiming the prevention of viral hepatitis in Brazil and may alert other countries in similar conditions. Disclosures: The following people have nothing to disclose: Rosangela Teixeira, Filipe M. Araújo, Emilio Tacrolimus (FK506) Suyama, Maria Aparecida M. Pereira, Geraldo Scarabelli, Soraia Z. Morais Introduction: In 2012, the American Board of Internal Medicine (ABIM) in collaboration with the American Association of the Study of Liver Diseases (AASLD) approved a competency-based Transplant Hepatology (TH) training pilot program. This program allows completion of both Gastroenterology (GI) and TH training in

three years. The institution of this pilot is a milestone towards a competency-based education model for training programs. The aim of this study was to identify the perceptions and beliefs of GI/Hepatology Division and Fellowship Program Directors on the combined GI/TH training pilot and competency-based education in GI fellowship. Methods: A 21 item survey was created to assess perceptions/beliefs about the 3-year combined GI/TH training pilot and the level of competency of graduates from the program. All current GI/TH Division and Fellowship Program Directors from AGCME-accredited programs were invited, via email to the Director, to anonymously complete the online survey (SurveyMonkeyTM). Results: A total of 1 1 6 participants completed the survey with a ~38% response rate.

e., three meeting abstracts

were included in the analysis for China and two in the analysis for Mexico). Seroprevalence using any type of HBsAg assay was allowed (complete criteria are described in Supporting Table 1). Fixed effect (FE) and random effects (RE) meta-analyses of HBsAg seroprevalence rates from studies that met the inclusion criteria were conducted to calculate country-specific pooled CHB prevalence rates. RE analysis, which assumes heterogeneity among surveys, was considered more appropriate based on the nature of the data: HBV was unevenly distributed and we expected different rates from different surveys carried out in different Z-IETD-FMK molecular weight populations in different locations at different times. FE analysis was conducted for comparison. Between-study heterogeneity was assessed for each country dataset using Cochran’s Q test

and the I2 statistic.14, 15 For most countries, data were insufficient for exploration of heterogeneity. Separate pooled rates were calculated for emigrants and for in-country populations for countries for which data were available, and results were compared using a Z test.15 Subgroup analyses were also done by decade of survey and by sex. For the 17 countries with at least 25 surveys, meta-regression analyses, based on the RE models using survey date as the covariate, Selleck Trametinib were done using Comprehensive Meta-Analysis software (Biostat, Englewood, NJ). For a few countries with low HBsAg seroprevalence rates (e.g., Etoposide datasheet Japan, Australia, New Zealand, Canada, and northern and western European countries), rates from large, population-based studies were used instead of meta-analysis. Study-quality assessment was done for only a subset of the data (i.e.,

Bangladesh, China, India, Iran, Korea, Pakistan, Philippines, Thailand, and Vietnam) to determine whether weighting based on study quality made a difference in the pooled prevalence rates. We developed a three-category scale (Supporting Table 2), scored each study, and calculated the pooled prevalence rates with and without the additional weighting factor, as described by Sutton et al.16 Flow of the systematic review is summarized by world region in Table 1. Results for individual countries are in Supporting Table 3. More than 17,500 articles were identified in PubMed searches; full text of 2,859 articles was assessed and data from 3,276 articles were entered into country-specific databases. In all, we found 1,373 articles reporting data meeting criteria for use in the meta-analyses. Many articles report data for more than one survey (e.g., pregnant women and military recruits) and these were entered separately. A total of 2,053 HBsAg seroprevalence surveys involving 18.6 million subjects were used in the meta-analyses (Table 2; Supporting Table 4).

Level C (possibly effective, ineffective, or harmful) rating requires at least 2 convincing class III studies. Adapted with permission from Brainin et al. Guidance for the preparation of neurological management guidelines by EFNS scientific task forces—revised recommendations 2004. Eur J Neurol 2004;11:577-581. “
“Migraine is a common primary headache disorder occurring predominantly in a young,

relatively healthy population. There is a growing literature on associations between migraine, especially migraine with aura, and ischemic stroke as well as other vascular events. U0126 Migraine as a risk factor for vascular disease and connections between migraine and endothelial, structural, and genetic risk are reviewed. There may be an interaction between endothelial dysfunction and cortical spreading depression affecting risk. Patient education and treatment of modifiable risk factors may decrease future vascular events. “
“(Headache 2011;51:860-868) Migraine is a common, often disabling disorder associated with a significant personal and societal burden. The presence of post-traumatic stress disorder (PTSD) may increase this disability substantially. Migraine and PTSD are both up to 3 times more common in women than in men. The divergence in prevalence rates of migraine and PTSD that occurs between the sexes after puberty suggests that gonadal hormones play an important role. In addition,

the preponderance of PTSD Belnacasan clinical trial in women may be related to their higher rates of interpersonal trauma, the most common cause of PTSD. However, recent data suggest that although the odds of PTSD are increased in both women and men with episodic migraine, PRKACG this association is stronger in men than women. In this paper, we examine the epidemiology of PTSD and migraine, with an emphasis on the known sex differences. We then discuss the neurobiological changes associated with PTSD, the current hypotheses for the mechanisms relating PTSD and migraine, and the treatment

implications of these findings. “
“Background.— In the absence of biological markers, the diagnosis of primary headache in epidemiological studies rests on clinical findings, as reported through ad-hoc interviews. Objectives.— The aim of this study was to validate a specially designed headache questionnaire to be administered by a physician for the diagnosis of primary headaches or of probable medication overuse headache in the general population according to the 2004 International Classification of Headache Disorders, 2nd edition (ICHD-II). Methods.— The questionnaire comprises 76 questions based on the ICHD-II diagnostic criteria for migraine (codes 1.1, 1.2.1, 1.2.2, 1.2.3, 1.5.1, and 1.6), tension-type headache (codes 2.1, 2.2, 2.3, and 2.4), primary stabbing headache (code 4.1), and probable medication-overuse headache (code 8.2.

Six such events have occurred in the Canary Islands but there have been no reported mass strandings in Hawai‘i. We assess the hypothesis that factors that influence the likelihood of strandings occurring and/or being detected differ between the Canary and main Hawaiian Islands, such that beaked whale stranding/detection probabilities will be lower in Hawai‘i. On an archipelago-wide basis, nearshore Vorinostat bathymetric

comparisons indicate that the Canaries have a greater proportion and a total greater amount of appropriate beaked whale habitat closer to shore, with a steeper slope. Hawaiian shorelines are more dominated by steep cliffs, human population density is much lower, and human population per kilometer of shoreline is 53% lower than in the Canaries. All of these factors suggest that there is a higher probability of a carcass washing onshore and being detected in the Canary Islands. It cannot be concluded that the lack of mass strandings in Hawai‘i is evidence of no impact. “
“Walleye pollock (Theragra chalcogramma) otoliths (n= 2,706) recovered check details from stomachs, small intestines, and colons of 43 northern fur seals (Callorhinus ursinus) were evaluated for size and wear by location in the digestive tract. Pollock fork length was regressed on otolith length after correction for erosion, and age

was estimated from the calculated body size. Age-1+ pollock otoliths (≥6.3-mm length) were concentrated in stomachs while age-0 otoliths (≤6.2-mm length) were concentrated in colons. Less than 10% of otoliths were found in the small intestines. Pollock age decreased with progression along seal gastrointestinal tracts. Otolith quality increased along gastrointestinal tracts in numbers ≥20, which was typical of age-0 otoliths recovered from colons. Otolith distribution by age and quality along gastrointestinal tracts suggests that small (≤12 cm) schooling prey are consumed in large volume and passed as a bolus rapidly through the digestive tract before significant erosion of bony remains

Celecoxib occurs; while larger prey are eaten in smaller volume and subjected to otolith erosion due to longer retention in the stomach. Our results illustrate the importance of multiple sampling strategies to comprehensively represent prey size in pinniped diet. “
“Stable carbon (δ13C) and nitrogen (δ15N) isotopes are used frequently to describe the trophic ecology of top marine mammal predators. Australian sea lions (Neophoca cinerea) are one of the world’s rarest otariid seals and exhibit the highest levels of natal site philopatry of any seal. We report the development of a screening technique to identify different foraging ecotypes and assess their relative frequencies in Australian sea lion breeding colonies using stable isotope ratios in pups.

The study was conducted in accordance with local Institutional Review Board regulations. We studied the following complications of ALI/ALF: hepatic encephalopathy, infection, systemic inflammatory response, renal failure, thrombosis, and bleeding. These complications were defined as follows: Hepatic encephalopathy was defined and graded according to West Haven criteria.[19]

Infection was defined as a positive urine culture, presence of a pulmonary FXR agonist infiltrate on chest X-ray consistent with infectious etiology, or a positive blood culture not felt to be a contaminant with a skin organism. More than one positive blood culture was required for bacteremia with commensal organisms. Systematic inflammatory response syndrome was defined according to established criteria[20]: white blood cell count >12 or <4 × 109 cells/L, temperature <36°C or >38°C, respiratory rate >20/minutes, and pulse >90 beats per minute. Renal failure was defined as persistent azotemia or oliguria despite rehydration requiring continuous veno-venous hemofiltration. Thrombosis was defined as occlusion of a native blood vessel or indwelling dialysis catheter. When occlusion of a native blood vessel was suspected on clinical grounds, these were confirmed by ultrasound or CT scanning. Bleeding

was defined as the presence of blood per naso-gastric tube, blood per rectum or endotracheal tube, or bleeding at the site of invasive procedure. Final outcomes of ALI/ALF were transplant-free survival, selleck chemicals llc orthotopic liver transplantation, or death. VWF antigen (VWF:Ag) levels were determined with an in-house

enzyme-linked immunosorbent assay (ELISA) assay using commercially available polyclonal antibodies against VWF (DAKO, Glostrup, Denmark). VWF ristocetin cofactor activity (VWF:RCo) was determined using the BC VWF-reagent (Siemens Healthcare Diagnostics) on a Behring Coagulation System (Siemens Healthcare Diagnostics). VWF:Ag and VWF:RCo levels of pooled normal plasma were set at 100% Casein kinase 1 and the values obtained in patient samples were expressed as a percentage of pooled normal plasma. VWF collagen binding activity was determined with an in-house ELISA assay as described.[8] The collagen-binding activity of pooled normal plasma was set at 100% and the activity measured in patient samples was expressed as a percentage of pooled normal plasma. VWF multimer analysis was performed by sodium dodecyl sulfate agarose gel electrophoresis followed by western blotting. The blots were incubated with rabbit anti-VWF antibody (DAKO) and goat anti-rabbit IRDye 800 CW (LI-COR Biosciences, Lincoln, NE). The first five bands were considered as low-molecular weight multimers, whereas other bands were considered as high molecular weight (HMW) multimers. The blots were scanned by the Odyssey Imager (Westburg, Leusden, The Netherlands) and were quantified by morphometric analysis using the ImageScope software package (Aperio, Vista, CA).