Barriers and facilitators in the referral pathways to low vision services from the perspective of patients and professionals: a qualitative study.

BACKGROUND: Underutilization of and lack of access to low vision services (LVS) has been reported internationally. The purpose of this study was to identify barriers and facilitators in LVS referral procedures and service delivery from both the perspective of people with visual impairment and professionals from different eye care providers in the Netherlands. METHODS: A qualitative study in the Netherlands was conducted. Barriers and facilitators were explored through semi structured interviews with older adults with macular degeneration, diabetic retinopathy and/or glaucoma (n = 14), and healthcare professionals including ophthalmologists and LVS professionals (n = 16). Framework analysis was used for analyzing the interviews with Atlas.ti software. RESULTS: According to both patients and professionals, facilitators in LVS access and utilization are having motivation, self-advocacy, high participation needs and social support, as well as being negatively impacted by the impairment. Both samples found having good communication skills and informing patients about LVS as a healthcare provider to facilitate access. A long patient-provider relationship and the Dutch healthcare system were also mentioned as facilitators. Professionals additionally found long disease duration and the presence of low vision optometric services in the ophthalmic practice to promote access. Barriers that were reported by patients and professionals are lack of motivation, self-advocacy and acceptance of the impairment in patients. In addition, having low participation needs as a patient, lack of information provision by providers and time constraints in the ophthalmic practice were mentioned as barriers. Professionals also reported lack of social support, short disease duration of patients, a short patient-provider relationship and lack of coordination of care in the ophthalmic practice to hinder access. CONCLUSIONS: Findings suggest that providers' lack of information provision about LVS, especially to patients who are less assertive, hamper referral to LVS. Providers should have attention for patients' LVS needs and actively inform them and their social network about LVS to facilitate access. Educating and training providers about how and when to address LVS may help to reduce barriers in the referral pathways. In addition, referral procedures may benefit from tools that make providers more aware of LVS.

Translation and content validity of the Dutch Impact of Vision Impairment questionnaire assessed by Three-Step Test-Interviewing.

BACKGROUND: Patients suffering from exsudative retinal diseases may experience severe central vision loss and this might have impact on their daily activities and quality of life. To measure the disabilities these patients may have, the use of the Impact of Vision Impairment Profile questionnaire is recommended. The aim of this study was to translate the original English 28-item Impact of Vision Impairment Profile (IVI) into the Dutch language and evaluate its comprehensibility, comprehensiveness and relevance as evidence of content validity. The translation process was performed using standardized methods. Content validity was assessed by cognitive debriefing using a Three-Step Test-Interview (TSTI) method for participants diagnosed with exudative retinal diseases. Step 1 and 2 focused on assessment of comprehensibility of items, step 3 on comprehensiveness and relevance. Audio-recorded qualitative data was analyzed using Atlas.ti. Data regarding comprehensibility problems was further categorized into item-specific problems and general problems. RESULTS: Few minor discrepancies in wording were found after translation. After conducting 12 cognitive interviews, data saturation was reached. All participants reported comprehensibility problems resulting from specific items, these were; sentence structure, vocabulary and formulation, influence of conditions or composite items and influence of comorbid disorders. Several general comprehensibility problems resulting from instructions or response categories were detected. The main general comprehensibility problem resulted from the layout of the Dutch-IVI. Most participants considered the included items as relevant and indicated that they covered the problems that occur due to vision impairment. CONCLUSIONS: Minor problems in the Dutch translation were detected and adjusted. The layout and instructions of the Dutch-IVI resulted in some comprehensibility problems. The Dutch-IVI appeared to be at risk of being interpreted as a generic patient reported outcome measure, instead of a disease-specific instrument, mainly due to the influence of co-morbidities. Adaptations should improve validity and reliability of the Dutch-IVI, however, cross-cultural comparisons may be at stake.

"Thinking a Lot" Among the Khwe of South Africa: A Key Idiom of Personal and Interpersonal Distress.

"Thinking too much", and variations such as "thinking a lot", are common idioms of distress across the world. The contextual meaning of this idiom of distress in particular localities remains largely unknown. This paper reports on a systematic study of the content and cause, consequences, and social response and coping related to the local terms |x'an n|a te and |eu-ca n|a te, both translated as "thinking a lot", and was part of a larger ethnographic study among the Khwe of South Africa. Semi-structured exploratory interviews with community members revealed that "thinking a lot" refers to a common experience of reflecting on personal and interpersonal problems. Consequences were described in emotional, psychological, social, behavioral, and physical effects. Coping strategies included social support, distraction, and religious practices. Our contextualized approach revealed meanings and experiences of "thinking a lot" that go beyond a psychological state or psychopathology. The common experience of "thinking a lot" is situated in socio-political, economic, and social context that reflect the marginalized and displaced position of the Khwe. We argue that "thinking a lot" and associated local meanings may vary across settings, may not necessarily indicate psychopathology, and should be understood in individual, interpersonal, community, and socio-political dimensions.

A review of barriers to using psychiatric advance directives in clinical practice.

Despite advocacy and demand for psychiatric advance directives (PADs), uptake and implementation in clinical practice is low. We examine why PAD implementation has been difficult globally by reviewing barriers in existing evidence. The review includes 30 studies, and identified 13 barriers, clustered into system level barriers, health professional level barriers, and service user level barriers. The considerable barriers to uptake and implementation hamper PAD use. We propose several potential strategies for overcoming some of the barriers. In order to realise these strategies, additional research is needed, particularly more field-based and operational research to understand processes and difficulties experienced in clinical practice.

Phosphoproteomic mass spectrometry profiling links Src family kinases to escape from HER2 tyrosine kinase inhibition.

Despite the initial effectiveness of the tyrosine kinase inhibitor lapatinib against HER2 gene-amplified breast cancers, most patients eventually relapse after treatment, implying that tumors acquire mechanisms of drug resistance. To discover these mechanisms, we generated six lapatinib-resistant HER2-overexpressing human breast cancer cell lines. In cells that grew in the presence of lapatinib, HER2 autophosphorylation was undetectable, whereas active phosphoinositide-3 kinase (PI3K)-Akt and mitogen-activated protein kinase (MAPK) were maintained. To identify networks maintaining these signaling pathways, we profiled the tyrosine phosphoproteome of sensitive and resistant cells using an immunoaffinity-enriched mass spectrometry method. We found increased phosphorylation of Src family kinases (SFKs) and putative Src substrates in several resistant cell lines. Treatment of these resistant cells with Src kinase inhibitors partially blocked PI3K-Akt signaling and restored lapatinib sensitivity. Further, SFK mRNA expression was upregulated in primary HER2+ tumors treated with lapatinib. Finally, the combination of lapatinib and the Src inhibitor AZD0530 was more effective than lapatinib alone at inhibiting pAkt and growth of established HER2-positive BT-474 xenografts in athymic mice. These data suggest that increased Src kinase activity is a mechanism of lapatinib resistance and support the combination of HER2 antagonists with Src inhibitors early in the treatment of HER2+ breast cancers in order to prevent or overcome resistance to HER2 inhibitors.

[Risk factors in adult periodontitis: polymorphism in the interleukin-1 gene family]. / Risicofactoren voor adulte parodontitis: polymorfismen in de interleukine-1 genfamilie.

Interleukin (IL)-1 alpha, IL-1 beta and IL-1 receptor antagonist (ra) play a major role in regulation of the inflammatory response in periodontal tissues. The aim of this study was to investigate the distribution of genetic variation in the IL-1 gene family among periodontitis patients and controls, taking into account smoking and microbiology as additional variables. There were 53 non-smoking and 52 smoking patients with severe adult periodontitis and 53 periodontal healthy controls genotyped for genetic variation in the IL-1 gene family. The presence of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans was established by culture techniques. A higher frequency of genotype+ (IL-1A*2 + IL-1B*2 + IL-1RN*2) was found in non-smoking periodontitis patients in whom P. gingivalis and A. actinomycetemcomitans could not be detected (42.1% vs. 11.3% in controls; p = 0.0068; or 5.7, 95% ci: 1.6-19.8). This data provide evidence that polymorphisms in genes of the IL-1 family are associated with severe adult periodontitis and may be a risk factor for severe periodontitis.

Polymorphisms of the interleukin-1 gene family, oral microbial pathogens, and smoking in adult periodontitis.

Interleukin (IL)-1alpha, IL-1beta, and IL-1ra contribute to regulation of the inflammatory response in periodontal tissues. We aimed to investigate the distribution of polymorphisms in the IL-1 gene family among periodontitis patients and controls, taking into account smoking and microbiology as additional variables. Fifty-three non-smoking and 52 smoking patients with severe adult periodontitis and 53 controls were genotyped for bi-allelic IL-1A(-889), IL-1B(-3954), and a penta-allelic 86-bp VNTR IL-1RN gene polymorphisms. The presence of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans was established by culture techniques. We found a higher frequency of allele 2 carriage in IL-1A, IL-1B, and IL-1RN in periodontitis patients who were non-smokers and in whom P. gingivalis and A. actinomycetemcomitans could not be detected (42.1% vs. 11.3% in controls; P = 0.0068; OR 5.7, 95% CI: 1.6-19.8). Our results provide evidence that polymorphisms in genes of the IL-1 family are associated with severe adult periodontitis in the absence of other risk factors tested in this patient population.

Simultaneous quantitation of N(2),3-ethenoguanine and 1,N(2)-ethenoguanine with an immunoaffinity/gas chromatography/high-resolution mass spectrometry assay.

We have previously described an immunoaffinity/gas chromatography/electron capture negative chemical ionization high-resolution mass spectrometry (IA/GC/ECNCI-HRMS) assay for quantitation of the promutagenic DNA adduct N(2),3-ethenoguanine (N(2),3-epsilonGua) in vivo. Here we present an expanded assay that allows simultaneous quantitation of its structural isomer, 1,N(2)-ethenoguanine (1,N(2)-epsilonGua), in the same DNA sample. 1,N(2)-epsilonGua and N(2),3-epsilonGua were purified together from hydrolyzed DNA using two immobilized polyclonal antibodies. GC/ECNCI-HRMS was used to quantitate the 3,5-bis(pentafluorobenzyl) (PFB) derivative of each adduct against an isotopically labeled analogue. Selected ion monitoring was used to detect the [M - 181](-) fragments of 3,5-(PFB)(2)-N(2),3-epsilonGua and 3,5-(PFB)(2)-[(13)C(4),(15)N(2)]-N(2),3-epsilonGua and the [M - 201](-) fragments of 3,5-(PFB)(2)-1,N(2)-epsilonGua and 3,5-(PFB)(2)-[(13)C(3)]-1,N(2)-epsilonGua. The demonstrated limits of quantitation in hydrolyzed DNA were 7.6 fmol of N(2),3-epsilonGua and 15 fmol of 1,N(2)-epsilonGua in approximately 250 microg of DNA, which corresponded to 5.0 N(2),3-epsilonGua and 8.7 1,N(2)-epsilonGua adducts/10(8) unmodified Gua bases, respectively. 1,N(2)-epsilonGua was found to be the predominant ethenoguanine adduct formed in reactions of lipid peroxidation products with DNA. The respective ratios of 1,N(2)-epsilonGua to N(2),3-epsilonGua were 5:1 and 38:1 when calf thymus DNA was treated with ethyl linoleate or 4-hydroxynonenal, respectively, under peroxidizing conditions. Only N(2),3-epsilonGua was detected in DNA treated with the vinyl chloride (VC) metabolite 2-chloroethylene oxide and in hepatocyte DNA from rats exposed to 1100 ppm VC for 4 weeks (6 h/day for 5 days/week). These data suggest that 1,N(2)-epsilonGua plays a minor role relative to N(2),3-epsilonGua in VC-induced carcinogenesis, but that 1,N(2)-epsilonGua may be formed to a larger extent from endogenous oxidative processes.

4-Hydroxy-2-nonenal and ethyl linoleate form N(2),3-ethenoguanine under peroxidizing conditions.

In these studies, we demonstrate that N(2),3-ethenoguanine (N(2), 3-epsilonGua) is formed from lipid peroxidation as well as other oxidative reactions. Ethyl linoleate (EtLA) or 4-hydroxy-2-nonenal (HNE) was reacted with dGuo in the presence of tert-butyl hydroperoxide (t-BuOOH) for 72 h at 50 degrees C. The resulting N(2), 3-epsilonGua was characterized by liquid chromatography/electrospray mass spectroscopy and by gas chromatography/high-resolution mass spectral (GC/HRMS) analysis of its pentafluorobenzyl derivative following immunoaffinity chromatography purification. The amounts of N(2),3-epsilonGua formed were 825 +/- 20 and 1720 +/- 50 N(2), 3-epsilonGua adducts/10(6) normal dGuo bases for EtLA and HNE, respectively, corresponding to 38- and 82-fold increases in the amount of N(2),3-epsilonGua compared to controls containing only t-BuOOH. Controls containing t-BuOOH but no lipid resulted in a >1000-fold increase in the level of N(2),3-epsilonGua over dGuo that was not subjected to incubation. EtLA and HNE, in the presence of t-BuOOH, were reacted with calf thymus DNA at 37 degrees C for 89 h. The amounts of N(2),3-epsilonGua formed in intact ctDNA were 114 +/- 32 and 52.9 +/- 16.7 N(2),3-epsilonGua adducts/10(6) normal dGuo bases for EtLA and HNE, respectively. These compared to 2.02 +/- 0. 17 and 2.05 +/- 0.47 N(2),3-epsilonGua adducts/10(6) normal dGuo bases in control DNA incubated with t-BuOOH, but no lipid. [(13)C(18)]EtLA was reacted with dGuo to determine the extent of direct alkylation by lipid peroxidation byproducts. These reactions resulted in a 89-93% level of incorporation of the (13)C label into N(2),3-epsilonGua when EtLA and dGuo were in equimolar concentrations, when EtLA was in 10-fold molar excess, and when deoxyribose (thymidine) was in 10-fold molar excess. Similar reactions with ctDNA resulted in an 86% level of incorporation of the (13)C label. These data demonstrate that N(2),3-epsilonGua is formed from EtLA and HNE under peroxidizing conditions by direct alkylation. The data also suggest, however, that N(2),3-epsilonGua is also formed by an alternative mechanism that involves some other oxidative reaction which remains unclear.

Increased p53 mutation load in noncancerous colon tissue from ulcerative colitis: a cancer-prone chronic inflammatory disease.

Ulcerative colitis (UC) is a chronic inflammatory disease that produces reactive oxygen and nitrogen species and increases the risk of colorectal cancer (CRC). The p53 tumor suppressor gene is frequently mutated in UC-associated dysplastic lesions and CRC. We are exploring the hypothesis that p53 mutations in the nontumorous colonic tissue in noncancerous UC cases indicate genetic damage from exposure to exogenous and endogenous carcinogens and may identify individuals at increased cancer risk. We are reporting, for the first time, the frequency of specific p53 mutated alleles in nontumorous colon tissue from donors either with or without UC by using a highly sensitive genotypic mutation assay. Higher p53 mutation frequencies of both G:C to A:T transitions at the CpG site of codon 248 and C:G to T:A transitions at codon 247 were observed in colon from UC cases when compared with normal adult controls (P = 0.001 and P = 0.001, respectively). In the UC cases, higher p53 codon 247 and 248 mutation frequencies were observed in the inflamed lesional regions when compared with the nonlesional regions of their colon (P < 0.001 and P = 0.001). The colonic nitric oxide synthase-2 activity was higher in UC cases than in non-UC adult controls (P = 0.02). Our data are consistent with the hypothesis that a higher frequency of p53 mutant cells can be generated under oxidative stress in people with UC. The increased frequency of specific p53 mutated alleles in noncancerous UC colon tissue may confer susceptibility to the development of CRC in an inflammatory microenvironment.

Immunoaffinity/gas chromatography/high-resolution mass spectrometry method for the detection of N(2),3-ethenoguanine.

Etheno adducts are formed after exposure to a number of carcinogens, including vinyl chloride, as well as endogenously as a result of lipid peroxidation. A sensitive and selective assay for N(2), 3-ethenoguanine (epsilonGua) was developed using immunoaffinity (IA) columns made with polyclonal antibodies to epsilonGua followed by gas chromatography/electron capture negative chemical ionization/high-resolution mass spectrometry (GC/ECNCI/HRMS) analysis of its pentafluorobenzyl derivative. These IA columns were specific for epsilonGua and did not bind guanine, deoxyguanosine, 1, N(6)-ethenoadenine, or 1,N(2)-ethenoguanine. The level of recovery of standards from the IA columns was 107 +/- 7% and throughout the entire method (using nucleoside enzymatic digestion) with or without DNA was 72 +/- 6%. Four different hydrolysis/digestion procedures were compared, nucleoside enzymatic (EZ), neutral thermal hydrolysis (NT), formic acid hydrolysis (FA), and HCl hydrolysis. All hydrolysis methods with subsequent IA chromatography produced linear standard curves with r(2) values of 0.999 or better. The level of epsilonGua in chloroethylene oxide-treated calf thymus DNA (CEO-ctDNA) was 38 +/- 2, 42 +/- 3, and 49 +/- 2 fmol of epsilonGua/microg of DNA using EZ, NT, and FA, respectively. These numbers remained consistent when the amount of DNA processed was doubled or tripled. These numbers were comparable to the previously published value of 55 +/- 8 fmol of epsilonGua/micrograms of DNA for the same DNA using HCl hydrolysis, cation exchange cleanup, and LC/MS analysis [Yen, T. Y., et al. (1996) J. Mass Spectrom. 31, 1271-1276]. Additionally, HCl hydrolysis of rat liver DNA from control and vinyl fluoride-exposed rats gave similar epsilonGua results when compared to those from enzymatic digestion using this method. This method gave a detection limit of 5 epsilonGua adducts/10(8) normal dGuo nucleosides in 150 micrograms of DNA using EZ and somewhat lower detection limits using NT and HCl hydrolysis. The method is more sensitive and selective than previously used methods for the quantitation of this adduct.

Antioxidant reactions of vitamin E in the perfused rat liver: product distribution and effect of dietary vitamin E supplementation.

We have investigated the relationship between vitamin E (alpha-tocopherol, TH) oxidation and antioxidant protection in a perfused rat liver model. Perfusion of a male Sprague-Dawley rat liver with 2 mM tert-butylhydroperoxide (t-BuOOH) for 10 min resulted in lipid peroxidation and metabolic changes reflecting oxidative stress. Mitochondria isolated from the liver exhibited increases in state 3 and state 4 respiration and a decline in the respiratory control ratio. In livers from rats given supplementary vitamin E in the diet, TH content was 7- to 10-fold higher than in controls and lipid peroxidation and metabolic changes induced by t-BuOOH were decreased. In mitochondria from these vitamin E-supplemented livers, the t-BuOOH-induced increase in state 4 respiration was reduced and the respiratory control ratio was maintained. In livers from unsupplemented rats, t-BuOOH induced oxidation of TH to alpha-tocopherolquinone, alpha-tocopherolhydroquinone, 2,3-epoxy-alpha-tocopherolquinone, and 5,6-epoxy-alpha-tocopherolquinone, as determined by gas chromatography-mass spectrometry analysis. Yields of these products were approximately doubled by treatment of samples with dilute acid, which indicated the presence of tocopherone and epoxytocopherone precursors. Oxidation of TH in vitamin E-supplemented livers yielded the same products and the relative extent of TH oxidation appeared similar to that in unsupplemented livers. In livers from both unsupplemented and vitamin E-supplemented animals, the distribution of oxidation products was similar in whole liver and isolated mitochondria. These data provide the first simultaneous documentation of TH antioxidant reactions and antioxidant effects in an intact organ system during oxidative stress.

Gas chromatography-mass spectrometry analysis of vitamin E and its oxidation products.

To facilitate studies of vitamin E ( alpha-tocopherol) antioxidant actions in tissues, we have developed a stable isotope dilution capillary gas chromatography-mass spectrometry assay for alpha-tocopherol and its three principal oxidation products, alpha-tocopherolquinone, 5,6-epoxy-alpha-tocopherolquinone, 2, 3-epoxy-alpha-tocopherolquinone, and for alpha-tocopherolhydroquinone, a reduction product of alpha-tocopherolquinone. Deuterium-labeled internal standards (5, 7-[2H3-methyl]-alpha-tocopherol, 2,6-[2H3-methyl]- alpha-tocopherolquinone, 2,6-[2H3-methyl]-5,6-epoxy- alpha-tocopherolquinone, 2,6-[2H3-methyl]-2,3-epoxy- alpha-tocopherolquinone, and 2-[2H3-methyl]- alpha-tocopherolhydroquinone are added to cell or tissue homogenates. The products then are extracted and converted to O-trimethylsilyl derivatives. Products are analyzed by capillary gas chromatography with on-column injection and detected by selected ion monitoring of characteristic fragment ions in electron ionization mode. Standard curves were linear from 25 fmol to 2 pmol for products and from 25 fmol to 4 pmol for alpha-tocopherol. The use of 2H6- and 2H3-internal standards for alpha-tocopherolquinone and alpha-tocopherolhydroquinone permits simultaneous analysis of both products despite possible redox interconversion during sample workup. alpha-Tocopherol oxidized in microsomes treated with azo-bis(amidinopropane HCl) was quantitatively accounted for as the epoxyquinones, alpha-tocopherolquinone, and alpha-tocopherolhydroquinone. However, over half of the oxidation products were present in microsomes as acid-labile tocopherone precursors. This method permits comprehensive assessment of vitamin E status in tissues and quantitative studies of vitamin E antioxidant actions and turnover.

Vitamin E oxidation in rat liver mitochondria.

Antioxidant reactions of alpha-tocopherol (vitamin E, alpha-TH) were studied by examining the fate of alpha-TH during oxidative challenge to mitochondrial membranes. Rat liver mitochondria were exposed to increasing concentrations of the water-soluble radical initiator 2,2'-azobis(2-amidinopropane) dihydrochoride (ABAP), and damage was assessed by monitoring mitochondrial respiration, alpha-TH oxidation, and lipid peroxidation. Significant lipid peroxidation was observed after 50% of the initial alpha-TH was depleted. Oxidative damage produced by ABAP-generated peroxyl radicals inhibited mitochondrial use of O2, as indicated by decreases in the respiratory control ratio and in state 3 and state 4 respiration. Rat liver mitochondria were supplemented with [14C]-alpha-TH by incubation of liver homogenate with [14C]-alpha-TH for 30 min at room temperature, followed by isolation of mitochondria by differential centrifugation. This supplementation resulted in a distribution of 83.7% and 14.3% of the added alpha-TH to the inner and outer mitochondrial membranes, respectively, which is similar to the distribution of endogenous alpha-TH. [14C]-alpha-TH-supplemented mitochondria then were treated with ABAP, and alpha-TH oxidation products were identified by radiochromatographic analysis of mitochondrial extracts. Products observed included alpha-tocopherolquinone, alpha-tocopherolquinone-2,3-oxide, and alpha-tocopherolquinone-5,6-oxide, which were identified by comparing HPLC retention and UV spectra to those of authentic standards. Product identities were verified by GC-MS of product O-trimethylsilyl derivatives. Another product, which was identified by HPLC, UV, and mass spectral analysis as 8a-(ethyldioxy)tocopherone, was found to be an artifact of sample workup and was shown to be derived from 8a-hydroperoxytocopherone, which was formed by alpha-TH oxidation in the mitochondria. These results indicate that alpha-TH antioxidant reactions in mitochondria are similar to those identified in homogeneous solutions and model liposomal systems.