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What makes immigration a salient issue among Latinos? We focus on immigration – one of the most pressing issues facing the United States (US) – and evaluate the factors that motivate immigration salience among Latinos over several election cycles. Although immigration policy has been linked with the Latino electorate over the period of our study due to high foreign-born rates and mixed-status families within this community, immigration policy has actually not been the dominant issue for the majority of Latino voters over this time period. Using survey responses from the 2008, 2012 and 2016 elections, we test multiple theories of issue salience by exploring social, political and individual determinants of policy salience among Latino voters. We find that in addition to nativity, consumption of ethnic media and group identity are associated with reporting immigration as a salient issue. These findings provide a valuable addition to literature of public opinion on immigration and the origins of policy issue salience among ethno-racial minorities in the US.

Appendectomy remains the standard of care for uncomplicated acute appendicitis despite several randomized clinical trials pointing to the safety and efficacy of nonoperative management of this disease. A meta-analysis of randomized clinical trials may contribute to the body of evidence and help surgeons select which patients may benefit from surgical and nonsurgical treatment.To assess the efficacy and safety of nonoperative management vs appendectomy for acute uncomplicated appendicitis.A systematic review was conducted using indexed sources (Embase and PubMed) to search for published randomized clinical trials in English comparing nonoperative management with appendectomy in adult patients presenting with uncomplicated acute appendicitis. To increase sensitivity, no limits were set for outcomes reported, sex, or year of publication. All nonrandomized or quasi-randomized trials were excluded, and validated primers were used.Among 1504 studies imported for screening, 805 were duplicates, and 595 were excluded for irrelevancy. A further 96 were excluded after full-text review, mainly owing to wrong study design or inclusion of pediatric populations. Eight studies met the inclusion criteria and were selected for the meta-analysis.Meta-extraction was conducted with independent extraction by multiple reviewers using the Covidence platform for systematic reviews and in accordance with PRISMA guidelines. Data were pooled by a random-effects model.Treatment success and major adverse effects at 30 days' follow-up.The main outcome (treatment success proportion at 30 days of follow-up) was not significantly different in the operative and nonoperative management cohorts (risk ratio [RR], 0.85; 95% CI, 0.66-1.11). Likewise, the percentage of major adverse effects was similar in both cohorts (RR, 0.72; 95% CI, 0.29-1.79). However, in the nonoperative management group, length of stay was significantly longer (RR, 1.48; 95% CI, 1.26-1.70), and a median cumulative incidence of 18% of recurrent appendicitis was observed.These results point to the general safety and efficacy of nonoperative management of uncomplicated acute appendicitis. However, this strategy may be associated with an increase in duration of hospital stay and a higher rate of recurrent appendicitis. This meta-analysis may help inform decision-making in nonoperative management of uncomplicated acute appendicitis.

Background: Demographic information related to phalangeal fractures that undergo simultaneous vascular repair, as well as their complication and reoperation profiles, remain incompletely understood. This study aimed to examine the patient and fracture characteristics influencing the outcomes after these injuries in a large Unites States adult patient cohort and to identify risk factors associated with unplanned reoperation of these fractures. Methods: A retrospective study was performed, identifying 54 phalangeal fractures in 48 patients; all fractures were also associated with vascular injuries requiring repair. Patients with digital amputations were excluded. A manual chart review was performed to collect epidemiologic, radiographic, and surgical outcome information. Results: The incidence of phalangeal fractures undergoing vascular repair was higher in the non-dominant hand, middle finger, proximal phalanx, and phalangeal shaft. Most (52.9%) fractures were due to occupational injury, with the most common mechanism being sharp injuries. More than half of the fractures had a nerve injury, and 13% required a vein graft for vascular repair. More than half of the fractures required at least one reoperation, most commonly due to “stiffness/tendon adhesion” (50%) and “nonunion or delayed union” (21.4%). In multivariable analysis, thumb (odds ratio [OR]: 35.1, P = .043) and index (OR: 14.0, P = .048) fingers’ fractures were found to be independently associated with unplanned reoperation. Conclusions: Phalangeal fractures requiring vascular repair occurred most often in the occupational setting and more than 50% required at least one unplanned reoperation. Injuries sustained in the thumb and index finger were more likely to undergo unplanned reoperation, which may guide initial treatment decision-making and postoperative follow-up.

Introduction The purpose of this study is to perform a cross-cultural adaptation and validation of the translated Patient-Rated Wrist Evaluation (PRWE) score exclusively for pathologies of the wrist. Materials and Methods A methodological study of cross-cultural validation of clinical scores was performed through a test–retest reliability analysis, internal consistency, response to change, and criterion validity assessment. Results The test was applied to 57 patients with 139 surveys. Stability evaluated through Lin's concordance correlation coefficient was 0.98, with 95% confidence interval (CI) = 0.97–0.99; Cronbach's alpha was > 0.91; the difference in score was 24.26 (standard deviation: 26.59); the standardized response mean was 0.912; the effect size was 0.924; the Spearman's coefficient between the differences of PRWE and DASH—Disabilities of the Arm, Shoulder, and Hand—scores was r = 0.899, with 95% CI = 0.811–0.947; Spearman's nonparametric correlation test between PRWE and DASH was 0.82, with 95% CI = 0.711–0.890. Conclusions We successfully validated the Spanish translation of the PRWE scale. It showed valid and reliable interpretation of functional status and response to treatment after distal radius fracture, for Colombian population. Level of Evidence This is a level II, methodological study for scale validation.

Previous research demonstrates that pandemics, including COVID-19, have disproportionate effects on communities of color, further exacerbating existing healthcare inequities. While increasing evidence points to the greater threat posed by COVID-19 to Latinx communities, less remains known about how identification as Latinx and migration status influence their perception of risk and harm. In this article, we use cross-sectional data from a large national probability sample to demonstrate a large positive association between ethnic identity and migration status and perceptions of harm from COVID-19 in the US. We find that individuals identifying as Hispanic/Latinx and first-generation immigrants report significantly greater risks of becoming infected by COVID-19 in the next three months, and dying from the virus if they do contract it. Further, subgroup analysis reveals that health risks are especially felt by individuals of Mexican descent, who represent the largest share of US Latinxs. Collectively, our results provide evidence about how the pandemic places increased stress on people from Latinx and immigrant communities relative to White non-Hispanic individuals in the US.

This study explores Latino perceptions of commonality and competition with African Americans across the country, focusing on the South. Using the Latino National Survey (LNS), we test the existing inter-group relation theories using an original measurement approach. With the creation of relative measures of commonality and competition of Latinos toward Blacks, we find that Latinos perceive co-ethnics as a greater source of competition than Blacks when our relative measure is used to interpret Latino perceptions of competition with African Americans. Moreover, our results suggest that Latinos in the South have similar perceptions of commonality to Blacks as Latinos more generally, across both approaches that measure perceptions of commonality. Most importantly, we find that when the relative competition measure is employed, Latinos who live in Southern states do in fact have higher perceptions of competition with Blacks than Latinos at large. These trends provide a valuable addition to the extant literature focused on inter-group relations by emphasizing that not only place and context matter, but also the way perceptions of competition and commonality are measured and operationalized.

While the literature on infectious disease outbreaks has examined the extent to which communication inequalities during public health emergencies exacerbate negative outcomes among disadvantaged individuals, the implications of ethnic media consumption among minority groups during these crises are underexplored. Making use of the first nationally representative survey of US Latinos (N = 1200) on the impact and reactions to COVID-19, this study examines the implications of Spanish-language news media consumption on source credibility and attitude formation during the COVID-19 pandemic among Latinos and immigrants from Latin America. Through a series of statistical analyses, this study finds that ethnic news consumption is strongly associated with trust in Spanish-language journalists, whereas mainstream media consumption is not associated with trust in English-language journalists. More importantly, this study finds that source credibility, particularly in Spanish-language journalists, matters for Latinos as it is associated with more positive assessments of state and local officials providing adequate information about COVID-19. This study illuminates the importance of non-traditional media among racial minorities, who account for almost 40% of the US population, and highlights the importance of shared backgrounds in source credibility among linguistically diverse groups in the United States during a public health crisis.

Social connectedness, sex, and intimacy are all factors associated with positive aging, facing individuals in society across the life course. Phenomenal technological developments in the 21st century have led to the increased use of smartphones, mobile apps, and dating apps for a myriad of services, and engagements. This paper focuses on two specific cohorts’ who have the opportunity to engage with dating apps, older adults and young citizens with life-limiting or life-threatening conditions, and highlights issues related to the intersection of technology, societal constructions of age, disability, and online dating.

Background Different bismuth quadruple therapies containing proton-pump inhibitors, bismuth salts, metronidazole, and a tetracycline have been recommended as third-line Helicobacter pylori eradication treatment after failure with clarithromycin and levofloxacin. Aim To evaluate the efficacy and safety of third-line treatments with bismuth, metronidazole, and either tetracycline or doxycycline. Methods Sub-study with Spanish data of the "European Registry on H pylori Management" (Hp-EuReg), international multicenter prospective non-interventional Registry of the routine clinical practice of gastroenterologists. After previous failure with clarithromycin- and levofloxacin-containing therapies, patients receiving a third-line regimen with 10/14-day bismuth salts, metronidazole, and either tetracycline (BQT-Tet) or doxycycline (BQT-Dox), or single capsule (BQT-three-in-one) were included. Data were registered at AEG-REDCap database. Univariate and multivariate analyses were performed. Results Four-hundred and fifty-four patients have been treated so far: 85 with BQT-Tet, 94 with BQT-Dox, and 275 with BQT-three-in-one. Average age was 53 years, 68% were women. Overall modified intention-to-treat and per-protocol eradication rates were 81% (BQT-Dox: 65%, BQT-Tet: 76%, BQT-three-in-one: 88%) and 82% (BQT-Dox: 66%, BQT-Tet: 77%, BQT-three-in-one: 88%), respectively. By logistic regression, higher eradication rates were associated with compliance (OR = 2.96; 95% CI = 1.01-8.84) and no prior metronidazole use (OR = 1.96; 95% CI = 1.15-3.33); BQT-three-in-one was superior to BQT-Dox (OR = 4.46; 95% CI = 2.51-8.27), and BQT-Tet was marginally superior to BQT-Dox (OR = 1.67; 95% CI = 0.85-3.29). Conclusion Third-line H pylori eradication with bismuth quadruple treatment (after failure with clarithromycin and levofloxacin) offers acceptable efficacy and safety. Highest efficacy was found in compliant patients and those taking 10-day BQT-three-in-one or 14-day BQT-Tet. Doxycycline seems to be less effective and therefore should not be recommended.

Introduction Esophageal atresia with or without tracheoesophageal fistula (EA/TEF) occurs approximately 1 in 3.500 live births representing the most common malformation of the upper digestive tract. Only half a century ago, EA/TEF was fatal among affected newborns suggesting that the steady birth prevalence might in parts be due to mutational de novo events in genes involved in foregut development. Methods To identify mutational de novo events in EA/TEF patients, we surveyed the exome of 30 case-parent trios. Identified and confirmed de novo variants were prioritized using in silico prediction tools. To investigate the embryonic role of genes harboring prioritized de novo variants we performed targeted analysis of mouse transcriptome data of esophageal tissue obtained at the embryonic day (E) E8.5, E12.5, and postnatal. Results In total we prioritized 14 novel de novo variants in 14 different genes (APOL2, EEF1D, CHD7, FANCB, GGT6, KIAA0556, NFX1, NPR2, PIGC, SLC5A2, TANC2, TRPS1, UBA3, and ZFHX3) and eight rare de novo variants in eight additional genes (CELSR1, CLP1, GPR133, HPS3, MTA3, PLEC, STAB1, and PPIP5K2). Through personal communication during the project, we identified an additional EA/TEF case-parent trio with a rare de novo variant in ZFHX3. In silico prediction analysis of the identified variants and comparative analysis of mouse transcriptome data of esophageal tissue obtained at E8.5, E12.5, and postnatal prioritized CHD7, TRPS1, and ZFHX3 as EA/TEF candidate genes. Re-sequencing of ZFHX3 in additional 192 EA/TEF patients did not identify further putative EA/TEF-associated variants. Conclusion Our study suggests that rare mutational de novo events in genes involved in foregut development contribute to the development of EA/TEF.

Different bismuth quadruple therapies containing proton-pump inhibitors, bismuth salts, metronidazole, and a tetracycline have been recommended as third-line Helicobacter pylori eradication treatment after failure with clarithromycin and levofloxacin.To evaluate the efficacy and safety of third-line treatments with bismuth, metronidazole, and either tetracycline or doxycycline.Sub-study with Spanish data of the "European Registry on H pylori Management" (Hp-EuReg), international multicenter prospective non-interventional Registry of the routine clinical practice of gastroenterologists. After previous failure with clarithromycin- and levofloxacin-containing therapies, patients receiving a third-line regimen with 10/14-day bismuth salts, metronidazole, and either tetracycline (BQT-Tet) or doxycycline (BQT-Dox), or single capsule (BQT-three-in-one) were included. Data were registered at AEG-REDCap database. Univariate and multivariate analyses were performed.Four-hundred and fifty-four patients have been treated so far: 85 with BQT-Tet, 94 with BQT-Dox, and 275 with BQT-three-in-one. Average age was 53 years, 68% were women. Overall modified intention-to-treat and per-protocol eradication rates were 81% (BQT-Dox: 65%, BQT-Tet: 76%, BQT-three-in-one: 88%) and 82% (BQT-Dox: 66%, BQT-Tet: 77%, BQT-three-in-one: 88%), respectively. By logistic regression, higher eradication rates were associated with compliance (OR = 2.96; 95% CI = 1.01-8.84) and no prior metronidazole use (OR = 1.96; 95% CI = 1.15-3.33); BQT-three-in-one was superior to BQT-Dox (OR = 4.46; 95% CI = 2.51-8.27), and BQT-Tet was marginally superior to BQT-Dox (OR = 1.67; 95% CI = 0.85-3.29).Third-line H pylori eradication with bismuth quadruple treatment (after failure with clarithromycin and levofloxacin) offers acceptable efficacy and safety. Highest efficacy was found in compliant patients and those taking 10-day BQT-three-in-one or 14-day BQT-Tet. Doxycycline seems to be less effective and therefore should not be recommended.

Introduction For parents of chronically ill children, the experiences of caregiving are challenged by increased demands and restrictions imposed by their child's disease. Therefore, this study aims to investigate the quality of life (QoL) in parents of children born with esophageal atresia (EA) and to explore associated factors. Materials and Methods Parents of children (2–17 years) with EA recruited from two German pediatric hospitals participated in this cross-sectional study about QoL in EA. Data on QoL, sociodemographic, and clinical characteristics were collected from parents and children. Parental QoL was assessed using the Short-Form 8 questionnaire, containing eight dimensions aggregated to a mental and physical health summary score which was compared with German representative population norms. Results Forty-nine families (47 mothers and 40 fathers) participated in the study. Compared with German population norms, both mothers and fathers showed significantly lower mental component score (MCS) but no differences in physical component score (PCS). Within the study sample, parents of younger children (2–7 years), severe EA, or high school/kindergarten absence had lower MCSs compared with those with older, less severe, and less absent children. Parental female gender was associated with lower MCS as well as lower family income. Conclusion Parents of children with EA reported lower mental health compared with the general population, especially mothers, and parents of young children, with severe EA, and a frequent school/kindergarten absence. This shows that parents may experience substantial emotional burden and highlights the need for psychosocial support for EA parents, especially in the first years.

This paper studies the dynamics of detention, deportation, and the criminalization of immigrants. We ground our analyses and discussion around the Illegal Immigration Reform and Immigrant Responsibility Act of 1996’s (IIRIRA’s) detention mandate, the role of special interest groups and federal policies. We argue that these special interest groups and major federal policies have come together to fuel the expansion of immigrant detention to unprecedented levels. Moreover, we aim to incite discussion on what this rapid growth in detention means for human rights, legislative representation and democracy in the United States. This study analyzes two main questions: What is the role of special interests in the criminalization of immigrants? And does the rapid increase in detention pose challenges or risks to democracy in the United States? Our study is grounded within the limited, yet growing literature on immigrant detention, government data, and “gray” literature produced by nonprofits and organizations working on immigration-related issues. We construct a unique dataset using this literature and congressional reports to assess what factors are associated with the rise of immigrant detention. A series of correlations and a time series regression analysis reveal that major restrictive federal immigration policies such as IIRIRA, along with the increasing federal immigration enforcement budget, have had a significant impact on immigrant detention rates. Based on these findings, we recommend three central policy actions. First, the paper recommends increased transparency and accountability on behalf of the Department of Homeland Security, Immigration and Customs Enforcement, and on lobbying expenditures from for-profit detention corporations. Second, it argues for the repeal of mandatory detention laws. These mandatory laws have led to the further criminalization and marginalization of undocumented immigrants. And lastly, it argues that repeal of the Congressional bed mandate would allow for the number of detainees to mirror actual detention needs, rather than providing an incentive to detain. However, we anticipate that the demand for beds will increase even more given the current administration’s push for the criminalization and increased arrests of undocumented individuals. The rhetoric used by the present administration further criminalizes immigrants.[1] [1] As reflected in the January 25, 2017 executive order from the White House: “Tens of thousands of removable aliens have been released into communities across the country, solely because their home countries refuse to accept their repatriation. Many of these aliens are criminals who have served time in our Federal, State, and local jails. The presence of such individuals in the United States, and the practices of foreign nations that refuse the repatriation of their nationals, are contrary to the human interest” (Enhancing Public Safety in the Interior of the United States, Exec. Order No. 13768, 82 Fed. Reg. 8799 [Jan. 25, 2017]).

Background The acronym VATER/VACTERL refers to the rare nonrandom association of the following component features (CF): vertebral defects (V), anorectal malformations (A), cardiac defects (C), tracheoesophageal fistula with or without esophageal atresia, renal malformations (R), and limb defects (L). Patients presenting with at least three CFs are diagnosed as having VATER/VACTERL association while patients presenting with only two CFs are diagnosed as having VATER/VACTERL-like phenotypes. Recently, rare causative copy number variations (CNVs) have been identified in patients with VATER/VACTERL association and VATER/VACTERL-like phenotypes. Methods To detect further causative CNVs we performed array based molecular karyotyping in 75 VATER/VACTERL and 40 VATER/VACTERL-like patients. Results Following the application of stringent filter criteria, we identified 13 microdeletions and seven microduplications in 20 unrelated patients all of which were absent in 1,307 healthy inhouse controls (n < 0.0008). Among these, microdeletion at 17q12 was confirmed to be de novo. Three microdeletions at 5q23.1, 16q23.3, 22q11.21, and one microduplication at 10q11.21 were all absent in the available parent. Microdeletion of chromosomal region 22q11.21 was previously found in VATER/VACTERL patients rendering it to be causative in our patient. The remaining 15 CNVs were inherited from a healthy parent. Conclusion In two of 115 patients' causative CNVs were found (2%). The remaining identified rare CNVs represent candidates for further evaluation. Rare inherited CNVs may constitute modifiers of, or contributors to, multifactorial VATER/VACTERL or VATER/VACTERL-like phenotypes. Birth Defects Research, 2017.© 2017 Wiley Periodicals, Inc.

The 2010 Patient Protection and Affordable Care Act (ACA) aimed to increase health insurance access for the over 47 million uninsured people in the U.S.A., among whom ethnoracial minorities had the...

While men and women make up a similar number of COVID-19 cases, and are equally likely to know someone who has become ill due to the virus, the gendered and systemic implications of immigration during public health emergencies among minority groups in the United States are empirically underexplored. Using the SOMOS COVID-19 Crisis National Latino Survey, we conduct a series of intersectional analyses to understand the extent to which personal experiences with COVID-19, gendered structural factors, and spillover effects of US immigration policies impact the mental health of US Latina/os during a public health emergency. The results show that among Latinas, knowing an undocumented immigrant and someone ill with COVID-19 increases the probability of reporting worse mental outcomes by 52 percent. Furthermore, being a woman increases the probability of reporting the highest level of mental health problems by 30 percent among Hispanic people who know someone with COVID-19 and an undocumented immigrant. These findings indicate that the effects of the COVID-19 outbreak among US Latinas and Latinos are entrenched in gendered and systemic inequalities.

Background Information on the epidemiology and susceptibility patterns of main pathogens causing severe acute diarrhea may help to reduce inappropriate antimicrobial use in emergency departments. Objectives We sought to investigate the micro-organisms causing severe acute diarrhea in patients requiring hospital admission by means of a commercial multiple polymerase chain reaction system. Methods Between November 2016 and October 2018 we studied 132 patients with acute diarrhea who required hospital admission at a 250-bed hospital in Spain. Demographic, clinical, analytical, and microbiological data were collected from the medical records. Stool samples were processed using a rapid commercial multiple polymerase chain reaction system (FilmArray Gastrointestinal Panel), stool culture, and standard microbiological procedures. Results The median age (range) of patients was 45.5 (0.1–92) years, and 50% were male; 46.2% presented with fever, 62.8% presented with vomiting, and 12.9% presented with rectal bleeding. At least 1 enteric pathogen was identified in 93 (70.4%) patients; 28 (21.2%) patients had >1 micro-organism. FilmArray Gastrointestinal Panel results were available in a median (range) of 1 (0–3) days. The micro-organisms most frequently identified were 24 cases of Campylobacter species, 20 cases of Clostridioides difficile producing toxin A or toxin B, 20 cases of Salmonella species, 12 cases of rotavirus, and 30 cases of different types of pathogenic Escherichia coli. Among the cases of C. difficile, 12 (60%) were community-acquired and 8 (40%) had an undetermined origin. Conclusion The FilmArray Gastrointestinal Panel system provides fast and reliable results and could be useful to select the most appropriate antimicrobial based on local susceptibilities until the results of the cultures are available.

Today’s health and social care systems are facing a challenge in how to effectively address caregiving for ageing populations facing cognitive disorders and frailty. Scholars and policy makers are now identifying a rise of “hidden form of care”, e.g. informal caregiving, as a phenomenon in support for ageing populations. Across Europe for instance, the rise in the older old adult population has led to a rapid expansion of the number of carers, both professional (formal) and informal. The latter, representing mostly family members caring for their loved ones, truly represents a “hidden form of care”. This can be a problem if formal and informal caregivers are not fully integrated into the healthcare continuum or are not given a systematic support to carry out caregiving in a relevant and safe way. There is currently no comprehensive European-wide legal framework and support mechanisms, in terms of training and education for this group. CARE Campus, an EIT Health programme within the Educational Campus Pillar, is a new model of collaboration between academic institutions, the private sector, and the public sector whose main aim is to support the development of a comprehensive training for formal and informal caregivers in Europe. The initial phase of the development encompasses nine (09) online training modules with a quality control process to ensure that the curriculum is evidence-based, compliant with the national and local regulations, and addresses the needs of caregivers across Europe. The objective is to support formal, informal, and family caregivers and reduce the burden on health care systems, whilst improving the quality of care for older adults.

Background Anorectal malformations (ARM) are rare abnormalities that occur in approximately 1 in 3000 live births with around 40% of patients presenting with isolated forms. Multiple familial cases reported, suggest underlying genetic factors that remain largely unknown. The recurrence in relatives is considered rare, however transmission rates of ARM by affected parents have never been determined before. The inheritance pattern of ARM was investigated in our database of patients with isolated ARM. Results Within our cohort of 327 patients with isolated ARM we identified eight adult patients from eight families who had in total 16 children with their healthy spouse. Of these ten had ARM, resulting in a recurrence risk of approximately one in two live births (10 of 16; 62%). From 226 families with 459 siblings we found two affected siblings in five families. Hence, the recurrence risk of ARM among siblings is approximately one in 92 live births (5 of 459; 1.0%). Conclusions Comparing the observed recurrence risk in our cohort with the prevalence in the general population, we see a 1500-fold increase in recurrence risk for offspring and a 32-fold increase if a sibling is affected. The recurrence risk of approximately 62% indicates an autosomal dominant mode of inheritance. Reliable figures on recurrence of ARM are becoming increasingly important since improved surgical techniques are able to maintain sexual function resulting in more offspring of patients with ARM. These data allow more precise counseling of families with ARM and support the need for genetic studies. Electronic supplementary material The online version of this article (10.1186/s13023-017-0729-7) contains supplementary material, which is available to authorized users.

N-glycosylation is a post-translational modification that plays a role in the trafficking and/or function of some membrane proteins. We have shown previously that N-glycosylation affected the function of some Kv1 voltage-gated potassium (K+) channels [Watanabe, Wang, Sutachan, Zhu, Recio-Pinto and Thornhill (2003) J. Physiol. (Cambridge, U.K.) 550, 51–66]. Kv1 channel S1–S2 linkers vary in length but their N-glycosylation sites are at similar relative positions from the S1 or S2 membrane domains. In the present study, by a scanning mutagenesis approach, we determined the allowed N-glycosylation sites on the Kv1.2 S1–S2 linker, which has 39 amino acids, by engineering N-glycosylation sites and assaying for glycosylation, using their sensitivity to glycosidases. The middle section of the linker (54% of linker) was glycosylated at every position, whereas both end sections (46% of linker) near the S1 or S2 membrane domains were not. These findings suggested that the middle section of the S1–S2 linker was accessible to the endoplasmic reticulum glycotransferase at every position and was in the extracellular aqueous phase, and presumably in a flexible conformation. We speculate that the S1–S2 linker is mostly a coiled-loop structure and that the strict relative position of native glycosylation sites on these linkers may be involved in the mechanism underlying the functional effects of glycosylation on some Kv1 K+ channels. The S3–S4 linker, with 16 amino acids and no N-glycosylation site, was not glycosylated when an N-glycosylation site was added. However, an extended linker, with an added N-linked site, was glycosylated, which suggested that the native linker was not glycosylated due to its short length. Thus other ion channels or membrane proteins may also have a high glycosylation potential on a linker but yet have similarly positioned native N-glycosylation sites among isoforms. This may imply that the native position of the N-glycosylation site may be important if the carbohydrate tree plays a role in the folding, stability, trafficking and/or function of the protein.

Kv1.4 and Kv1.1 potassium channels are expressed in brain as mature glycoproteins that are trans-Golgi glycosylated. When expressed in cell lines these homomers had very different trans-Golgi glycosylation efficiencies and cell surface expression levels with Kv1.4 > Kv1.1 for both parameters (Zhu, J., Watanabe, I., Gomez, B., and Thornhill, W. B. (2001) J. Biol. Chem. 276, 39419–39427). This previous study identified determinants in the outer pore region of Kv1.4 and Kv1.1 that positively and negatively, respectively, affected these events when expressed as homomers. Here we investigated which subunit exhibited positive or negative effects on these processes when expressed as heteromers. Kv1.4/Kv1.1 heteromers, by coexpression or expression as tandem-linked heteromers, were expressed on the cell surface at ∼20-fold lower levels versus Kv1.4 homomers but they were trans-Golgi glycosylated. The lower Kv1.4/Kv1.1 expression level was not rescued by Kvβ 2.1 subunits. Thus Kv1.1 inhibited high cell surface expression and partially retained the heteromer in the endoplasmic reticulum, whereas Kv1.4 stimulated trans-Golgi glycosylation. The subunit determinants and cellular events responsible for these differences were investigated. In a Kv1.4/Kv1.1 heteromer, the Kv1.1 pore was a major negative determinant, and it inhibited high cell surface expression because it induced high partial endoplasmic reticulum retention and it decreased protein stability. Other Kv1.1 regions also inhibited high surface expression of heteromers. The Kv1.1 C terminus induced partial Golgi retention and contributed to a decreased protein stability, whereas the Kv1.1 N terminus contributed to only a decreased protein stability. Thus a neuron may regulate its cell surface K+ channel protein levels by different Kv1 subfamily homomeric and heteromeric combinations that affect intracellular retention characteristics and protein stability.

Different classes of Kv1 potassium channels have different trafficking patterns despite having very similar amino acid sequences. Two amino acids responsible for these differences have been identified in the outer pore turret region of Kv1.1 and Kv1.4. Here we tested a series of substitutions at these two determinants on Kv1.4. All P506 substitutions tested resulted in a significant decrease in surface protein, total protein, and protein half-life, indicating that proline is required at 506 to stabilize protein conformation and increase trafficking to the cell surface. All K533 substitutions tested had no effect on total protein, suggesting that the lysine at 533 is not important for maintaining Kv1.4 protein conformation. However, a basic or long polar amino acid, such as K, R, or Q, at this position favored high surface protein and efficient trafficking of Kv1.4, whereas an acidic or short amino acid, such as D, E, S, L, N, or H, at this position induced partial high endoplasmic reticulum-retention. This intracellular retention was not due to protein misfolding. We propose that these four prolines and four lysines in a Kv1.4 homotetramer might provide a binding site for a putative endoplasmic reticulum-export molecule to ensure high cell surface protein expression of the channel.

Kv1.1 and Kv1.4 potassium channels are expressed as mature glycosylated proteins in brain, whereas they exhibited striking differences in degree of trans-Golgi glycosylation conversion and high cell surface expression when they were transiently expressed as homomers in cell lines. Kv1.4 exhibited a 70% trans-Golgi glycosylation conversion, whereas Kv1.1 showed none, and Kv1.4 exhibited a approximately 20-fold higher cell surface expression level as compared with Kv1.1. Chimeras between Kv1.4 and Kv1.1 and site-directed mutants were constructed to identify amino acid determinants that affected these processes. Truncating the cytoplasmic C terminus of Kv1.4 inhibited its trans-Golgi glycosylation and high cell surface expression (as shown by Li, D., Takimoto, K., and Levitan, E. S. (2000) J. Biol. Chem. 275, 11597-11602), whereas truncating this region on Kv1.1 did not affect either of these events, indicating that its C terminus is not a negative determinant for these processes. Exchanging the C terminus between these channels showed that there are other regions of the protein that exert a positive or negative effect on these processes. Chimeric constructs between Kv1.4 and Kv1.1 identified their outer pore regions as major positive and negative determinants, respectively, for both trans-Golgi glycosylation and cell surface expression. Site-directed mutagenesis identified a number of amino acids in the pore region that are involved in these processes. These data suggest that there are multiple positive and negative determinants on both Kv1.4 and Kv1.1 that affect channel folding, trans-Golgi glycosylation conversion, and cell surface expression.

Background/Purpose: Mesenchymal hamartoma (MH) of the liver is thought to develop from the ductal plates of the prenatal liver. This immunohistochemical study was performed to gain insight into the pathophysiology of its development. Methods: Specimens from four MHs with adjacent liver, in one case from a biopsy and from the resected lesion after 6 years follow-up, were investigated with immunostaining on cryostat sections with antibodies against cytokeratins, vimentin, desmin and alpha-actin, as well as von Willebrand factor (factor VIII), fibroblast growth factor (FGF) receptors, FGF-1 (acidic FGF), FGF-2 (basic FGF), and the proliferation-associated Ki67 antigen. Results: Fibrous tissue of MH stained positive not only for vimentin, but also for desmin and alpha-actin, whereas cytokeratins and factor VIII showed specific staining in biliary cysts and endothelial cells, respectively. All mesenchymal cells expressed proteins of the FGF receptor family. Although FGF-1 was only scarcely detectable, there was an accumulation of FGF-2 in borderline areas of liver to MH. Multiple Ki67-positive mesenchymal cells could be identified in these regions in all three MHs. However, we could not detect any proliferative activity in the MHs after follow-up. Conclusions: The proliferative process in MH is still active during early childhood. FGF-2 may have a role in promoting this process. The positivity for desmin and alpha-actin of the lesions suggests that fat-storing (Ito) cells of the immature liver may be involved in the development of MH.

We have shown previously that truncating all of the variable cytoplasmic C-terminus of Kv1.1 potassium channels to G421stop had only a small inhibitory effect on their cell surface conductance density levels and cell surface protein levels. Here we investigated the role of a highly conserved cytoplasmic C-terminal charged region of five amino acids (HRETE) of the S6 transmembrane domain in the protein and conductance expression of Kv1.1, Kv1.2, and Kv1.4 channels. For Kv1.1 we found that E420stop, T419stop, and E418stop showed cell surface conductance densities and cell surface protein levels similar to full length control, whereas R417stop and H416stop exhibited essentially no conductance but their surface protein levels were similar to full length control. A bulky non-negatively charged hydrophilic amino acid at position 417 appeared to be critical for wild type gating of Kv1.1 because R417K and R417Q rescued conductance levels whereas R417A or R417E did not. The R417A mutation in the full length Kv1.1 also exhibited surface protein levels similar to control but it did not exhibit significant conductance. In contrast, mutation of the equivalent arginine to alanine in full length Kv1.2 and Kv1.4 appeared to have little or no effect on channel conductance but rather decreased cell surface protein levels by inducing partial high ER retention. These findings are consistent with the notion that the arginine amino acid in the HRETE region plays a different role in affecting conductance levels or cell surface protein levels of very closely related Kv1 potassium channels.

Kv1.4 and Kv1.1 potassium channel homomers have been shown to exhibit different intracellular trafficking programmes and cell-surface expression levels in cell lines: a determinant in the pore region of Kv1.4 and Kv1.1 [Zhu, Watanabe, Gomez and Thornhill (2001) J. Biol. Chem. 276, 39419–39427] and a cytoplasmic C-terminal VXXSL determinant on Kv1.4 [Li, Takimoto and Levitan (2000) J. Biol. Chem. 275, 11597–11602] have been described, which affected trafficking and cell-surface expression levels. In the present study, we examined whether trafficking pore determinants influenced any cytoplasmic C-terminal trafficking determinant. We found that removal of VXXSL from a Kv1.4 chimaera that contained the pore of Kv1.1 did not affect cell-surface trafficking. Therefore removal of the C-terminal VXXSL of Kv1.4 inhibited protein surface levels only in the presence of the Kv1.4 pore. In contrast, truncating the cytoplasmic C-terminus of Kv1.1 or truncating a Kv1.1 chimaera with the pore of Kv1.4, had little effect on surface protein levels. Furthermore, the subregion of the Kv1.4 pore trafficking determinant that was required for the inhibitory effect of VXXSL removal was mapped to a threonine residue in the deep pore region. Therefore the Kv1.4 pore determinant affected the trafficking and cell-surface levels directed by the C-terminal VXXSL determinant. Different Kv1 trafficking programmes would affect cell-surface expression levels either positively or negatively and also cell signalling. Cells may use differential trafficking programmes of membrane proteins as a post-translational mechanism to regulate surface protein levels and cell function.