Development of a canine artificial colonic mucus model for drug diffusion studies.

Colonic mucus is a key factor in the colonic environment because it may affect drug absorption. Due to the similarity of human and canine gastrointestinal physiology, dogs are an established preclinical species for the assessment of controlled release formulations. Here we report the development of an artificial colonic mucus model to mimic the native canine one. In vitro models of the canine colonic environment can provide insights for early stages of drug development and contribute to the implementation of the 3Rs (refinement, reduction, and replacement) of animal usage in the drug development process. Our artificial colonic mucus could predict diffusion trends observed in native mucus and was successfully implemented in microscopic and macroscopic assays to study macromolecular permeation through the mucus. The traditional Transwell set up was optimized with the addition of a nylon filter to ensure homogenous representation of the mucus barrier in vitro. In conclusion, the canine artificial colonic mucus can be used to study drug permeation across the mucus and its flexibility allows its use in various set ups depending on the nature of the compound under investigation and equipment availability.

Social Insurance Literacy of Dutch Workers Receiving Disability Benefits and its Associations with Socio-Economic Characteristics.

Purpose This study explores the concept social insurance literacy (SIL) and corresponding questionnaire (SILQ) among workers receiving disability benefits and the comprehensibility of the social security institute (SSI), and examines associations with socio-economic characteristics. Methods 1753 panel members of the Dutch SSI were approached to complete the SILQ-NL37. This measure was based on the original SILQ. The SILQ-NL37 contains domains for obtaining, understanding and acting upon information for both individual SIL and system comprehensibility. A higher score means better SIL or comprehensibility. Data on age, gender, education, living situation, Dutch skills and time receiving disability benefits were also collected. With k-means clustering, groups with adequate and limited SIL were created. Associations with socio-economic characteristics were examined with independent t-tests and linear regression analyses for both the total scores and within domain scores. Cronbach α and Spearman rho's indicated measurement properties were good to acceptable for the SILQ-NL37. Results Thirty-five percent of the 567 participants were in the group with limited SIL. Higher individual SILQ-NL37 scores were associated with having a partner (p = 0.018) and northeastern living region (p = 0.031). Higher scores for obtaining (p = 0.041) and understanding (p = 0.049) information were associated with female sex, and for acting on information with younger age (p = 0.020). People with limited Dutch skills (p = 0.063) and a partner (p = 0.085) rated system comprehensibility higher. Conclusions According to the SILQ-NL37 scores, about 35% of the panel members have limited ability to obtain, understand and act upon social insurance systems information. Limited SIL is associated with several socio-economic factors. Future researches should study the concept in a more representative sample, and in different countries and social insurance contexts.

An updated investigation of the multidimensional structure of language lateralization in left- and right-handed adults: a test-retest functional transcranial Doppler sonography study with six language tasks.

A previous study we reported in this journal suggested that left and right-handers may differ in their patterns of lateralization for different language tasks (Woodhead et al. 2019 R. Soc. Open Sci. 6, 181801. (doi:10.1098/rsos.181801)). However, it had too few left-handers (N = 7) to reach firm conclusions. For this update paper, further participants were added to the sample to create separate groups of left- (N = 31) and right-handers (N = 43). Two hypotheses were tested: (1) that lateralization would be weaker at the group level in left-than right-handers; and (2) that left-handers would show weaker covariance in lateralization between tasks, supporting a two-factor model. All participants performed the same protocol as in our previous paper: lateralization was measured using functional transcranial Doppler sonography during six different language tasks, on two separate testing sessions. The results supported hypothesis 1, with significant differences in laterality between groups for four out of six tasks. For hypothesis 2, structural equation modelling showed that there was stronger evidence for a two-factor model in left than right-handers; furthermore, examination of the factor loadings suggested that the pattern of laterality across tasks may also differ between handedness groups. These results expand on what is known about the differences in laterality between left- and right-handers.

An influenza A (H3N2) virus outbreak in the Kingdom of Cambodia during the COVID-19 pandemic of 2020.

BACKGROUND: Global influenza virus circulation decreased during the COVID-19 pandemic, possibly due to widespread community mitigation measures. Cambodia eased some COVID-19 mitigation measures in June and July 2020. On 20 August a cluster of respiratory illnesses occurred among residents of a pagoda, including people who tested positive for influenza A but none who were positive for SARS-CoV-2. METHODS: A response team was deployed on 25 August 2020. People with influenza-like illness (ILI) were asked questions regarding demographics, illness, personal prevention measures, and residential arrangements. Respiratory swabs were tested for influenza and SARS-Cov-2 by real-time reverse transcription PCR, and viruses were sequenced. Sentinel surveillance data were analyzed to assess recent trends in influenza circulation in the community. RESULTS: Influenza A (H3N2) viruses were identified during sentinel surveillance in Cambodia in July 2020 prior to the reported pagoda outbreak. Among the 362 pagoda residents, 73 (20.2%) ILI cases were identified and 40 were tested, where 33/40 (82.5%) confirmed positive for influenza A (H3N2). All 40 were negative for SARS-CoV-2. Among the 73 residents with ILI, none were vaccinated against influenza, 47 (64%) clustered in 3/8 sleeping quarters, 20 (27%) reported often wearing a mask, 27 (36%) reported often washing hands, and 11 (15%) reported practicing social distancing. All viruses clustered within clade 3c2.A1 close to strains circulating in Australia in 2020. CONCLUSIONS: Circulation of influenza viruses began in the community following the relaxation of national COVID-19 mitigation measures, and prior to the outbreak in a pagoda with limited social distancing. Continued surveillance and influenza vaccination are required to limit the impact of influenza globally.

Combined volar and dorsal plating vs. volar plating of distal radius fractures. A single-center study of 105 cases.

Despite recent advances in the surgical management of distal radius fractures (DRFs), the optimal treatment remains controversial as different fixation methods often have similar clinical functional and radiographic outcomes. The objective of this study was to assess the differences in outcomes 1 year postoperatively between volar plating and combined plating for DRFs. In a retrospective cohort study, we evaluated 105 consecutive patients operated with either a volar locking plate or combined dorsal and volar plating. The primary outcome was wrist range of motion (ROM). Secondary outcome measures included hand grip strength, visual analog scale (VAS) pain scores, radiographic examination and patient-related outcome measures. Patients treated with combined plating had significantly inferior wrist flexion, extension and ulnar deviation. The radiographic Batra score 1 year postoperatively was similar for both groups. The PRWE (patient-rated wrist evaluation) score was 16 for the volar plating group and 14 for the combined plating group. The QuickDASH (Quick disabilities of the arm, shoulder and hand) score was 9 for the volar plating group and 16 for the combined plating group. VAS pain scores were 0 at rest and 2 during activity for both groups. Grip strength was similar between the two groups. Hardware removal was done in 18/78 patients for the combined plating group and 1/27 for the volar plate group. Two patients operated with combined plating had tendon ruptures. Our findings indicate that both methods can yield satisfactory clinical and radiographic outcomes. However, combined plating resulted in inferior wrist ROM and substantially higher frequency of hardware removal. The potential advantages of combined plating in stabilizing a comminuted DRF must be balanced by the potential drawbacks such as inferior wrist ROM and higher frequency of hardware removal.

Warming weather changes the chemical composition of oat hulls.

The current threats of climate change are driving attention away from the petrochemical industry towards more sustainable and bio-based production processes for fuels and speciality chemicals. These processes require suitable low-cost starting material. One potential material assessed here is the oat hull. Its overall chemical composition has so far not been fully characterized. Furthermore, it is not known how it is affected by extreme weather events. Oat hulls (Kerstin and Galant varieties) grown during 'normal' weather years (2016 and 2017) are compared to the harvest of the warmer and drier year (2018). Standard methods for determination of plant chemical composition, with focus on carbohydrate composition, are utilized. Oat hulls grown in 'normal' weather conditions (2017) are rich in lignocellulose (84%), consisting of 35% hemicellulose, 25% lignin and 23% cellulose. Arabinoxylan was found to be the major biopolymer (32%). However, this composition is greatly influenced by weather variations during the oat growth phase. A lignocellulose reduction of 25% was recorded in the warmer and drier 2018 harvest. Additionally, a 6.6-fold increase in starch content, a four-fold increase in protein content and a 60% decrease in phenolic content was noted. Due to its high lignocellulose composition, with an exceptionally large hemicellulose fraction, the chemical composition of oat hulls is unique among agricultural by-products. However, this characteristic is significantly reduced when grown in warmer and drier weather, which could compromise its suitability for use in a successful biorefinery.

Tularaemia in European Brown Hares (Lepus europaeus) and Mountain Hares (Lepus timidus) Characterized by Histopathology and Immunohistochemistry: Organ Lesions and Suggestions of Routes of Infection and Shedding.

Tularaemia is an emerging zoonotic infectious disease caused by the bacterium Francisella tularensis. In Sweden, hares are considered to be key species in the epidemiology of tularaemia. The aim of this study was to characterize the pathology of natural tularaemia infection in European brown hares (EBHs) (Lepus europaeus) and mountain hares (MHs) (Lepus timidus) in Sweden, in order to better understand the presentation of disease and the routes of infection, body dissemination and shedding of F. tularensis. During 2000-2013, 49 EBHs and 37 MHs were diagnosed with tularaemia. Enlargement of the spleen was seen in 80% of EBHs and 62% of MHs. Necrosis was often obvious in the bone marrow, liver, lung and spleen, but 30% of the hares had no lesions or minimal gross lesions. On microscopical examination of tissues from 27 EBHs and three MHs, necrosis was seen in the majority of samples of liver, spleen, bone marrow, lymph node and adrenal glands and was common in the lungs and brain meninges. Immunohistochemistry for Francisella spp. detected bacteria in association with necrosis and inflammation. In several cases, Francisella spp. were also found inside blood vessels, in the renal pelvis, in lactating mammary glands, in bronchioles and in the skin, associated with tick bites. Using quantitative polymerase chain reaction, two genotypes of F. tularensis subsp. holarctica were found; canSNP group B.6, all belonging to subgroup B.7, and canSNP group B.12. There were no differences in pathology between the genotypes. Our results indicate that the urinary tract and mammary glands are important routes for the shedding of F. tularensis. Hunters may not be aware of the risks of contracting tularaemia while handling hares, since infected hares do not always show noticeable gross lesions.

Increased risk of influenza among vaccinated adults who are obese.

BACKGROUND: Influenza infects 5-15% of the global population each year, and obesity has been shown to be an independent risk factor for increased influenza-related complications including hospitalization and death. However, the risk of developing influenza or influenza-like illness (ILI) in a vaccinated obese adult population has not been addressed. OBJECTIVE: This study evaluated whether obesity was associated with increased risk of influenza and ILI among vaccinated adults. SUBJECTS AND METHODS: During the 2013-2014 and 2014-2015 influenza seasons, we recruited 1042 subjects to a prospective observational study of trivalent inactivated influenza vaccine (IIV3) in adults. A total of 1022 subjects completed the study. Assessments of relative risk for laboratory confirmed influenza and ILI were determined based on body mass index. Seroconversion and seroprotection rates were determined using prevaccination and 26-35 days post vaccination serum samples. Recruitment criteria for this study were adults 18 years of age and older receiving the seasonal trivalent inactivated influenza vaccine (IIV3) for the years 2013-2014 and 2014-2015. Exclusion criteria were immunosuppressive diseases, use of immunomodulatory or immunosuppressive drugs, acute febrile illness, history of Guillain-Barre syndrome, use of theophylline preparations or use of warfarin. RESULTS: Among obese, 9.8% had either confirmed influenza or influenza-like-illness compared with 5.1% of healthy weight participants. Compared with vaccinated healthy weight, obese participants had double the risk of developing influenza or ILI (relative risk=2.01, 95% CI 1.12, 3.60, P=0.020). Seroconversion or seroprotection rates were not different between healthy weight and obese adults with influenza or ILI. CONCLUSIONS: Despite robust serological responses, vaccinated obese adults are twice as likely to develop influenza and ILI compared with healthy weight adults. This finding challenges the current standard for correlates of protection, suggesting use of antibody titers to determine vaccine effectiveness in an obese population may provide misleading information.

High and novel genetic diversity of Francisella tularensis in Germany and indication of environmental persistence.

In Germany tularemia is a re-emerging zoonotic disease. Therefore, we investigated wild animals and environmental water samples for the presence and phylogenetic diversity of Francisella tularensis in the poorly studied Berlin/Brandenburg region. The phylogenomic analysis of three isolates from wild animals revealed three new subclades within the phylogenetic tree of F. tularensis [B.71 from a raccoon dog (Nyctereutes procyonoides); B.74 from a red fox (Vulpes vulpes), and B.75 from a Eurasian beaver (Castor fiber albicus)]. The results from histological, PCR, and genomic investigations on the dead beaver showed that the animal suffered from a systemic infection. Indications were found that the bacteria were released from the beaver carcass into the surrounding environment. We demonstrated unexpectedly high and novel phylogenetic diversity of F. tularensis in Germany and the fact that the bacteria persist in the environment for at least one climatic season. These findings support a broader host species diversity than previously known regarding Germany. Our data further support the assumption derived from previous serological studies of an underestimated frequency of occurrence of the pathogen in the environment and in wild animals. F. tularensis was isolated from animal species not previously reported as natural hosts in Germany.

Scaffolding of a bacterial genome using MinION nanopore sequencing.

Second generation sequencing has revolutionized genomic studies. However, most genomes contain repeated DNA elements that are longer than the read lengths achievable with typical sequencers, so the genomic order of several generated contigs cannot be easily resolved. A new generation of sequencers offering substantially longer reads is emerging, notably the Pacific Biosciences (PacBio) RS II system and the MinION system, released in early 2014 by Oxford Nanopore Technologies through an early access program. The latter has highly advantageous portability and sequences samples by measuring changes in ionic current when single-stranded DNA molecules are translocated through nanopores. We show that the MinION system produces long reads with high mapability that can be used for scaffolding bacterial genomes, despite currently producing substantially higher error rates than PacBio reads. With further development we anticipate that MinION will be useful not only for assembling genomes, but also for rapid detection of organisms, potentially in the field.

Memory Brought to Mind. Five-Year Follow-Up of Case Finding and Intervention of Dementia in a Swedish Primary Health Care District.

BACKGROUND: In an urban/rural primary health care (PHC) district a five-year integrated project of early detection and management of cognitive disorders was made in collaboration between home care (HC) and family practice services using a single-item case-finding and intervention approach. OBJECTIVES: To assess feasibility, outcome and morbidity over a 5-year period. DESIGN, SETTING AND PARTICIPANT: In autumn 2008, the question "Have you experienced memory difficulties or been told of them by family members?" was mailed to all eligible persons > 75 years of age (n=367) in the urban/rural Vålberg HC and PHC district (population = 5073). 320 (= 87%; 184 no and 136 yes) responded and 117 yes-responders came for further examination. In the follow-up, all diagnoses up till November 2013 were collected and compared anonymously in both yes and no answerers. RESULTS: 114 completed examination. 29 showed low risk of cognitive impairment, 39 moderate and 46 high. Definitive diagnosis was obtained in 34 of the latter: 10 cognitive impairment, 16 Alzheimer's disease, 5 non-specific, 2 vascular and 1 alcoholic dementia. During follow-up no further dementia diagnoses occurred in the low, two in the moderate, and none in the high-risk group, versus 12 in the no responders. Age and mortality were significantly higher in the high-risk group. Co-morbidity was very frequent but did not differ between the groups. CONCLUSIONS: Population response and compliance were excellent; the single-item direct question approach gave workable results with in particular high negative predictive power persisting over the five-year follow-up period, and can be applied in early case-finding, prevention and intervention of cognitive impairment in an integrated local HC, PHC and Hospital setting.

Breast cancer during follow-up and progression - A population based cohort on new cancers and changed biology.

BACKGROUND: Emerging data indicate an important role for biopsies of clinically/radiologically defined breast cancer 'recurrences'. The present study investigates tumour related events (relapses, other malignancies, benign conditions) after a primary breast cancer diagnosis. PATIENTS AND METHODS: The cohort includes 2102 women, representing all patients, with primary invasive breast cancer during 2000-2011 in the county of Värmland, Sweden. A comparative analysis of oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and proliferation (Ki67) between the primary tumour and the relapse was performed and related to outcome. RESULTS: With a mean follow-up time of 4.8 years, 1060 out of 2102 patients have had a biopsy taken after the initial breast cancer diagnosis demonstrating 177 recurrences, 93 other malignancies (colorectal, lung, skin), 40 cancer in situ (skin, breast) and 857 benign lesions. Approximately 70% (177 out of 245) of all cases of relapsed breast cancer underwent a biopsy during this time period. For patients with recurrences, ER (n=127), PR (n=101), HER2 (n=73) and Ki67 (n=55) status in both primary tumour and the corresponding relapse were determined. The discordance of receptor status was 14.2%, 39.6%, 9.6% and 36.3%, respectively. Loss of ER or PR in the relapse resulted in a significant increased risk of death (hazard ratio (HR) 3.62; 95% confidence interval (CI), 1.65-7.94) and (HR 2.34; 95% CI, 1.01-5.47) compared with patients with stable ER or PR positive tumours. The proportion of patients losing ER was bigger in the group treated with endocrine therapy alone or in combination with chemotherapy, 16.7% and 13.3%, respectively, compared with the group treated with chemotherapy alone or that which received no treatment 4.3% and 7.7%, respectively. CONCLUSION: Discordance of biomarkers between the primary tumour and the corresponding relapse was seen in 10-40% of the patients, adjuvant therapies seem to drive clonal selections. Patients with tumours losing ER or PR during progression have worse survival compared with patients with retained receptor expression.

Clonal alteration of breast cancer receptors between primary ductal carcinoma in situ (DCIS) and corresponding local events.

BACKGROUND: Emerging data propose biomarker alteration due to clonal selection between the primary invasive breast cancer and corresponding metastases. In addition, impact on survival has been demonstrated. The present study investigates the relationship between the oestrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) between primary ductal carcinoma in situ (DCIS) and intra-individually matched ipsilateral event. MATERIALS AND METHODS: The cohort includes 1504 patients, diagnosed with a primary DCIS between 1986 and 2004. Of the 274 patients who developed a local relapse, 135 developed a new in situ carcinoma and 139 an invasive cancer up to 31st December 2011. ER and PR were identified by immunohistochemistry (IHC) and HER2 by silver-enhanced in situ hybridisation (SISH) as well as IHC. RESULTS: ER (n=112), PR (n=113) and HER2 (n=114) status from both the primary DCIS and the corresponding relapse were assessed and were demonstrated to be discordant in 15.1%, 29.2% and 10.5% respectively. The receptor conversion was both from negative to positive and from positive to negative with no general pattern being seen in spite of sub-dividing into in situ relapse and invasive relapse. However, primary DCIS was HER2 positive in 40.3% whereas in situ and invasive relapses were HER2 positive in 42.9% and 34.5% respectively. CONCLUSIONS: Receptor conversion for ER, PR and HER2 status occurred between primary DCIS and corresponding local relapse in 10-30%. This study could not confirm that HER2 overexpression in primary DCIS had any impact on tumour progression to invasive cancer which has been proposed.

Isoflurane affects the cytoskeleton but not survival, proliferation, or synaptogenic properties of rat astrocytes in vitro.

BACKGROUND: More than half of the cells in the brain are glia and yet the impact of general anaesthetics on these cells is largely unexamined. We hypothesized that astroglia, which are strongly implicated in neuronal well-being and synapse formation and function, are vulnerable to adverse effects of isoflurane. METHODS: Cultured rat astrocytes were treated with 1.4% isoflurane in air or air alone for 4 h. Viability, proliferation, and cytoskeleton were assessed by colorimetric assay, immunocytochemistry, or a migration assay at the end of treatment or 2 days later. Also, primary rat cortical neurones were treated for 4 days with conditioned medium from control [astrocyte-conditioned media (ACM)], or isoflurane-exposed astrocytes (Iso-ACM) and synaptic puncta were assessed by synapsin 1 and PSD-95 immunostaining. RESULTS: By several measures, isoflurane did not kill astrocytes. Nor, based on incorporation of a thymidine analogue, did it inhibit proliferation. Isoflurane had no effect on F-actin but reduced expression of α-tubulin and glial fibrillary acidic protein both during exposure (P<0.05 and P<0.001, respectively) and 2 days later (P<0.01), but did not impair astrocyte motility. ACM increased formation of PSD-95 but not synapsin 1 positive puncta in neuronal cultures, and Iso-ACM was equally effective. CONCLUSIONS: Isoflurane decreased expression of microtubule and intermediate filament proteins in astrocytes in vitro, but did not affect their viability, proliferation, motility, and ability to support synapses.

Obesity is associated with impaired immune response to influenza vaccination in humans.

BACKGROUND: Obesity is an independent risk factor for morbidity and mortality from pandemic influenza H1N1. Influenza is a significant public health threat, killing an estimated 250,000-500,000 people worldwide each year. More than one in ten of the world's adult population is obese and more than two-thirds of the US adult population is overweight or obese. No studies have compared humoral or cellular immune responses to influenza vaccination in healthy weight, overweight and obese populations despite clear public health importance. OBJECTIVE: The study employed a convenience sample to determine the antibody response to the 2009-2010 inactivated trivalent influenza vaccine (TIV) in healthy weight, overweight and obese participants at 1 and 12 months post vaccination. In addition, activation of CD8⁺ T cells and expression of interferon-γ and granzyme B were measured in influenza-stimulated peripheral blood mononuclear cell (PBMC) cultures. RESULTS: Body mass index (BMI) correlated positively with higher initial fold increase in IgG antibodies detected by enzyme-linked immunosorbent assay to TIV, confirmed by HAI antibody in a subset study. However, 12 months post vaccination, higher BMI was associated with a greater decline in influenza antibody titers. PBMCs challenged ex vivo with vaccine strain virus, demonstrated that obese individuals had decreased CD8⁺ T-cell activation and decreased expression of functional proteins compared with healthy weight individuals. CONCLUSION: These results suggest obesity may impair the ability to mount a protective immune response to influenza virus.

Assessment of enzymatic prodrug stability in human, dog and simulated intestinal fluids.

The aim of this study was to determine the stability of three ester prodrugs, chloramphenicol succinate, enalapril and candesartan cilexetil, in human proximal small intestinal fluid (HIF), dog proximal small intestinal fluids (DIF) and simulated intestinal fluid (FaSSIF), with the addition of pancreatin. The total protein content in the proximal jejunal fluids was determined in HIF and DIF, respectively. Candesartan cilexetil was significantly degraded in HIF (initial t(1/2(0-5 min))=5.4 ± 0.5 min) and in DIF (initial t(1/2(0-5 min))=5.7 ± 0.1 min), while chloramphenicol succinate and enalapril were stable in both media. The degradation of candesartan cilexetil was shown to be mediated by enzymes following Michaelis-Menten enzyme kinetics and was inhibited by addition of esterase inhibitors. The enzymatic capacity reflected by V(max) was 4-fold higher in DIF than in HIF and correlated to its 2-fold higher protein concentration. The degradation of candesartan cilexetil in the FaSSIF-pancreatin solution was slower (t(1/2)=207 ± 34 min) than the degradation in both HIF and DIF. Changing the pH to the enzyme optima or increasing the amount of pancreatin, increased the degradation rate of candesartan cilexetil, but not in the magnitude as in HIF. As a result, two in vitro models, based on in vivo intestinal fluids, were developed using candesartan cilexetil as a model drug. The DIF seems to be a reasonably good model for HIF, although the degradation capacity seems to be somewhat higher, possibly due to the higher enzyme concentration in DIF. Future investigations will develop novel enzymatic based in vitro models for rapid assessment and biopharmaceutical screening tools for prodrugs.

Integrated HPLC-MS and (1)H-NMR spectroscopic studies on acyl migration reaction kinetics of model drug ester glucuronides.

Acyl glucuronides (AGs) are common, chemically reactive metabolites of acidic xenobiotics. Concerns about the potential of this class of conjugate to cause toxicity in man require efficient methods for the determination of reactivity, and this is commonly done by measuring transacylation kinetics. High-performance liquid chromatography-mass spectrometry (HPLC-MS) and nuclear magnetic resonance (NMR) spectroscopy were applied to the kinetic analysis of AG isomerization and hydrolysis for the 1-beta-O-AGs of ibufenac, (R)- and (S)-ibuprofen, and an alpha,alpha-dimethylated ibuprofen analogue. Each AG was incubated in either aqueous buffer at pH 7.4 or human plasma at 37 degrees C. Aliquots of these samples, taken throughout the reaction time course, were analysed by HPLC-MS and (1)H-NMR spectroscopy and the results compared. For identification of the AGs incubated in pH 7.4 buffer and for analysis of kinetic rates, (1)H-NMR spectroscopy generally gave the most complete set of data, but for human plasma the use of (1)H-NMR spectroscopy was impractical and HPLC-MS was more suitable. HPLC-MS was more sensitive than (1)H-NMR spectroscopy, but the lack of suitable stable-isotope labelled internal standards, together with differences in response between glucuronides and aglycones, made quantification problematic. Using HPLC-MS a specific 1-beta-O-AG-related ion at m/z 193 (the glucuronate fragment) was noted enabling selective determination of these isomers. In buffer, transacylation reactions predominated, with relatively little hydrolysis to the free aglycone observed. In human plasma incubations the observed rates of reaction were much faster than for buffer, and hydrolysis to the free aglycone was the major route. These results illustrate the strengths and weaknesses of each analytical approach for this class of analyte.

An investigation of serum concentration of apoM as a potential MODY3 marker using a novel ELISA.

OBJECTIVE: To investigate the fitness of serum apolipoprotein M (apoM) concentration as a marker for maturity-onset diabetes of the young 3 (MODY3). STUDY DESIGN AND SUBJECTS: This study consisted of two parts. A family study included 71 carriers of the P291fsinsC mutation in hepatocyte nuclear factor-1alpha (HNF-1alpha) from the Finnish Botnia study, 53 of whom were diabetic, and 75 matched family controls. A second, case-control study included 24 MODY3 patients, 17 healthy MODY3 mutation carriers, 11 MODY1 patients, 18 type 2 diabetes patients and 19 healthy control individuals. Subjects in the case-control study were recruited from the Botnia study or the Clinic of Endocrinology, Malmö University Hospital. Serum apoM levels were measured using a novel ELISA based on two monoclonal apoM antibodies. RESULTS: In the family study, mean serum apoM was 10% lower in female carriers of the P291fsinsC mutation compared to the family controls (P = 0.0058), a difference which remained significant after adjustment for diabetes status. There was no observed difference between groups for men. In the case-control study, no significant difference in apoM concentration was observed between MODY3 and type 2 diabetes patients, neither before nor after adjustment for total cholesterol. CONCLUSIONS: Female carriers of the P291fsinsC mutation in HNF-1alpha displayed slightly lower apoM serum levels. This difference is too small for apoM to be reliably employed as a biomarker for HNF-1alpha mutation status.

Large differences in levodopa dose requirement in Parkinson's disease: men use higher doses than women.

BACKGROUND AND PURPOSE: The characteristics of levodopa dosing are not well described in the literature. The aims were to investigate the use of levodopa in a nationwide Swedish survey and to study the characteristics of low-dose and high-dose patients with Parkinson's disease (PD) in a university hospital. METHODS: Patients with >or= 1 and >or= 2 purchases of levodopa during 2007 were selected from the prescribed drug register. Daily levodopa doses were estimated. Records of 504 patients with PD who visited the neurology clinic at Uppsala University Hospital during 2006-2007 were examined to select a low-dose group (< or = 400 mg levodopa daily, n = 21) and a high-dose group (>or= 1200 mg daily, n = 26) with at least 5 years of PD duration. RESULTS: In total, 33 534 levodopa users with > or = 1 levodopa purchase were found. Daily levodopa dose range was large; median daily dose was 465 mg for men and 395 mg for women (P < 0.0001). Almost half (46%) of the patients used < 400 mg levodopa daily. Significantly, more men were treated with doses >or= 1200 mg daily. Dose and age correlated negatively (P < 0.0001). Patients with high dose at 5 years PD duration continuously increased their dosage the following years, whereas low-dose patients did not. The occurrence of dyskinesias was about the same in both groups despite the large difference in levodopa dose. CONCLUSIONS: We conclude that the levodopa requirement in PD ranges considerably, and that men use higher levodopa dose than women. Levodopa requirement is constant during the progression of the disease in low-dose patients but increases in high-dose patients.