In situ production and secretion of proteins endow therapeutic benefit against psoriasiform dermatitis and melanoma.

Genetic medicines have the potential to treat various diseases; however, certain ailments including inflammatory diseases and cancer would benefit from control over extracellular localization of therapeutic proteins. A critical gap therefore remains the need to develop and incorporate methodologies that allow for posttranslational control over expression dynamics, localization, and stability of nucleic acid-generated protein therapeutics. To address this, we explored how the body's endogenous machinery controls protein localization through signal peptides (SPs), including how these motifs could be incorporated modularly into therapeutics. SPs serve as a virtual zip code for mRNA transcripts that direct the cell where to send completed proteins within the cell and the body. Utilizing this signaling biology, we incorporated secretory SP sequences upstream of mRNA transcripts coding for reporter, natural, and therapeutic proteins to induce secretion of the proteins into systemic circulation. SP sequences generated secretion of various engineered proteins into the bloodstream following intravenous, intramuscular, and subcutaneous SP mRNA delivery by lipid, polymer, and ionizable phospholipid delivery carriers. SP-engineered etanercept/TNF-α inhibitor proteins demonstrated therapeutic efficacy in an imiquimod-induced psoriasis model by reducing hyperkeratosis and inflammation. An SP-engineered anti-PD-L1 construct mediated mRNA encoded proteins with longer serum half-lives that reduced tumor burden and extended survival in MC38 and B16F10 cancer models. The modular nature of SP platform should enable intracellular and extracellular localization control of various functional proteins for diverse therapeutic applications.

GLUT3 promotes macrophage signaling and function via RAS-mediated endocytosis in atopic dermatitis and wound healing.

The facilitative GLUT1 and GLUT3 hexose transporters are expressed abundantly in macrophages, but whether they have distinct functions remains unclear. We confirmed that GLUT1 expression increased after M1 polarization stimuli and found that GLUT3 expression increased after M2 stimulation in macrophages. Conditional deletion of Glut3 (LysM-Cre Glut3fl/fl) impaired M2 polarization of bone marrow-derived macrophages. Alternatively activated macrophages from the skin of patients with atopic dermatitis showed increased GLUT3 expression, and a calcipotriol-induced model of atopic dermatitis was rescued in LysM-Cre Glut3fl/fl mice. M2-like macrophages expressed GLUT3 in human wound tissues as assessed by transcriptomics and costaining, and GLUT3 expression was significantly decreased in nonhealing, compared with healing, diabetic foot ulcers. In an excisional wound healing model, LysM-Cre Glut3fl/fl mice showed significantly impaired M2 macrophage polarization and delayed wound healing. GLUT3 promoted IL-4/STAT6 signaling, independently of its glucose transport activity. Unlike plasma membrane-localized GLUT1, GLUT3 was localized primarily to endosomes and was required for the efficient endocytosis of IL-4Rα subunits. GLUT3 interacted directly with GTP-bound RAS in vitro and in vivo through its intracytoplasmic loop domain, and this interaction was required for efficient STAT6 activation and M2 polarization. PAK activation and macropinocytosis were also impaired without GLUT3, suggesting broader roles for GLUT3 in the regulation of endocytosis. Thus, GLUT3 is required for efficient alternative macrophage polarization and function, through a glucose transport-independent, RAS-mediated role in the regulation of endocytosis and IL-4/STAT6 activation.

Red blood cells as glucose carriers to the human brain: Modulation of cerebral activity by erythrocyte exchange transfusion in Glut1 deficiency (G1D).

Red blood cells circulating through the brain are briefly but closely apposed to the capillary endothelium. We hypothesized that this contact provides a nearly direct pathway for metabolic substrate transfer to neural cells that complements the better characterized plasma to endothelium transfer. While brain function is considered independent of normal fluctuations in blood glucose concentration, this is not borne out by persons with glucose transporter I (GLUT1) deficiency (G1D). In them, encephalopathy is often ameliorated by meal or carbohydrate administration, and this enabled us to test our hypothesis: Since red blood cells contain glucose, and since the red cells of G1D individuals are also deficient in GLUT1, replacing them with normal donor cells via exchange transfusion could augment erythrocyte to neural cell glucose transport via mass action in the setting of unaltered erythrocyte count or plasma glucose abundance. This motivated us to perform red blood cell exchange in 3 G1D persons. There were rapid, favorable and unprecedented changes in cognitive, electroencephalographic and quality-of-life measures. The hypothesized transfer mechanism was further substantiated by in vitro measurement of direct erythrocyte to endothelial cell glucose flux. The results also indicate that the adult intellect is capable of significant enhancement without deliberate practice. ClinicalTrials.gov registration: NCT04137692 https://clinicaltrials.gov/ct2/show/NCT04137692.

Characterization of ALTO-encoding circular RNAs expressed by Merkel cell polyomavirus and trichodysplasia spinulosa polyomavirus.

Circular RNAs (circRNAs) are a conserved class of RNAs with diverse functions, including serving as messenger RNAs that are translated into peptides. Here we describe circular RNAs generated by human polyomaviruses (HPyVs), some of which encode variants of the previously described alternative large T antigen open reading frame (ALTO) protein. Circular ALTO RNAs (circALTOs) can be detected in virus positive Merkel cell carcinoma (VP-MCC) cell lines and tumor samples. CircALTOs are stable, predominantly located in the cytoplasm, and N6-methyladenosine (m6A) modified. The translation of MCPyV circALTOs into ALTO protein is negatively regulated by MCPyV-generated miRNAs in cultured cells. MCPyV ALTO expression increases transcription from some recombinant promoters in vitro and upregulates the expression of multiple genes previously implicated in MCPyV pathogenesis. MCPyV circALTOs are enriched in exosomes derived from VP-MCC lines and circALTO-transfected 293T cells, and purified exosomes can mediate ALTO expression and transcriptional activation in MCPyV-negative cells. The related trichodysplasia spinulosa polyomavirus (TSPyV) also expresses a circALTO that can be detected in infected tissues and produces ALTO protein in cultured cells. Thus, human polyomavirus circRNAs are expressed in human tumors and infected tissues and express proteins that have the potential to modulate the infectious and tumorigenic properties of these viruses.

Caregiving experiences of Korean family caregivers of cancer patients: An integrative literature review.

OBJECTIVE: The purpose of this study was to describe the experiences of Korean family caregivers of patients with cancer by reviewing recent literature. METHODS: Five electronic databases were searched-MEDLINE, the Cumulative Index to Nursing and Allied Health Literature (CINAHL), and PubMed for English literature, and Korean Information Service System (KISS), and Research Information Sharing Service (RISS) for Korean literature for articles published from January 2010 to March 2020 using the key words cancer, caregiver, and Korean. Twenty-six articles met the inclusion criteria and remained in the final review. RESULTS: No intervention study was found and most of studies were quantitative without theoretical/conceptual framework. All the studies were conducted with Koreans living in Korea. No previous study has been conducted with Koreans living in the U.S. or other countries. Most studies focused on caregivers' quality of life, burden, unmet needs, and resilience/adaptation/post-traumatic growth. CONCLUSIONS: Comprehensive intervention studies focused on improving quality of life, burden, and adaptation to their complex roles as caregivers in the context of Korean culture would be helpful. Further research is needed in examining the caregiver-patient dyad interactions longitudinally to understand the dynamic complicated processes of caregiving.

Perceptions of illness severity in adults with drug-resistant temporal lobe epilepsy.

OBJECTIVE: The purpose of this study was to explore how subjective perceptions of illness severity were described by a sample of participants with drug-resistant epilepsy (DRE) who were considering surgery. METHODS: A qualitative methodology, constructivist grounded theory, guided all aspects of the study. Data were collected via 51 semi-structured interviews with 35 adults in our multiethnic sample. At interview, the 20 women (57%) and 15 men (43%) ranged in age from 18 to 68 years (mean = 35.6 years) and had lived with epilepsy for an average of 15.4 y (range = 2-44 years). RESULTS: A grounded theory with four interrelated categories was developed to reflect the process by which participants arrived at an explanation of illness severity. Illness severity for participants evolved as participants reflected upon the burdensome impact of uncontrolled seizures on self and others. Epilepsy, when compared with other chronic conditions, was described as less serious, and participants imagined that other peoples' seizures were comparatively worse than their own. Illness severity was not uppermost in participants' minds but emerged as a concept that was both relative and linked to social burden. Perceptions of overall disease severity expanded upon determinants of seizure severity to offer a more complete explanation of what patients themselves did about longstanding, uncontrolled epilepsy. CONCLUSIONS: Perceptions of illness severity played a vital role in treatment decision-making with the potential to impact the illness trajectory. How to measure components of illness severity represents a new challenge for outcomes research in DRE.

Adaptation and validation of the health belief model scale for colorectal cancer screening.

Korea has the second highest incidence of colorectal cancer in the world. Instruments that are culturally and contextually sensitive, as well as valid and reliable, for determining health beliefs regarding colorectal cancer screening are essential for obtaining accurate information. The purpose of this study was to adapt and validate the health belief model scale for Koreans regarding colorectal cancer and fecal occult blood test utilization. Individual and cognitive interviews (also known as cognitive debriefing) with 33 Koreans, expert reviews with seven nursing practitioners and professors, and a pilot test with 18 Koreans were conducted to make the existing health belief model scale culturally and contextually sensitive. Subsequently, a cross-sectional survey with 728 Koreans aged >50 years was conducted. Exploratory and confirmatory factor analyses of the construct validity and internal consistency reliability supported the adapted health belief model scale. The adapted and validated health belief model scale in this study could contribute to the assessment of health beliefs regarding the fecal occult blood test among Koreans with a greater degree of accuracy with respect to Korean culture and context.

Assessment of circularized E7 RNA, GLUT1, and PD-L1 in anal squamous cell carcinoma.

Anal squamous cell carcinoma (ASCC) is a rare, potentially fatal malignancy primarily caused by high-risk human papillomaviruses (HPV). The prognostic implication of programmed death-ligand 1 (PD-L1) expression remains controversial, and glucose transporter 1 (GLUT1) expression has never been examined in ASCC. Covalently closed circular RNAs have recently been shown to be widespread in cancers and are proposed to be biomarkers. We discovered HPV16 expresses a circular E7 RNA (circE7) which has not been assessed as a potential biomarker. A retrospective, translational case series at UT Southwestern was conducted to analyze PD-L1, GLUT1, HPV-ISH, and HPV circE7 in relation to the clinical features and overall survival of patients with ASCC. Twenty-two (22) subjects were included in the study. Improved overall survival was predicted by basaloid histology ( p = 0.013), PD-L1 expression ( p = 0.08), and HPV-ISH positivity ( p & 0.001), but not GLUT1 expression. High levels of circE7 by quantitative RT-PCR predicted improved overall survival in ASCC ( p = 0.023) and analysis of The Cancer Genome Atlas sequencing from HPV-positive head and neck cancer and cervical cancer suggested high circE7 marked improved survival in 875 subjects ( p = 0.074). While our study suggests that circE7 levels correlate with improved survival in ASCC, larger, prospective studies are necessary to confirm the potential role of circE7 as a biomarker.

Trichodysplasia spinulosa in a child: Identification of trichodysplasia spinulosa-associated polyomavirus in skin, serum, and urine.

A 6-year-old girl with a history of chronic immunosuppression following small bowel and colon transplantation for tufting enteropathy presented with a diffuse, facial-predominant eruption composed of pink-to-skin-colored papules with central white dystrophic spicules. Histology from a punch biopsy and polymerase chain reaction (PCR) from plucked spicules confirmed a diagnosis of trichodysplasia spinulosa (TS). Additional molecular studies identified several strains of the trichodysplasia spinulosa-associated polyomavirus infecting multiple tissues of the patient, confirming the systemic nature of trichodysplasia spinulosa infections.

Transforming activity of an oncoprotein-encoding circular RNA from human papillomavirus.

Single-stranded circular RNAs (circRNAs), generated through 'backsplicing', occur more extensively than initially anticipated. The possible functions of the vast majority of circRNAs remain unknown. Virus-derived circRNAs have recently been described in gamma-herpesviruses. We report that oncogenic human papillomaviruses (HPVs) generate circRNAs, some of which encompass the E7 oncogene (circE7). HPV16 circE7 is detectable by both inverse RT-PCR and northern blotting of HPV16-transformed cells. CircE7 is N6-methyladenosine (m6A) modified, preferentially localized to the cytoplasm, associated with polysomes, and translated to produce E7 oncoprotein. Specific disruption of circE7 in CaSki cervical carcinoma cells reduces E7 protein levels and inhibits cancer cell growth both in vitro and in tumor xenografts. CircE7 is present in TCGA RNA-Seq data from HPV-positive cancers and in cell lines with only episomal HPVs. These results provide evidence that virus-derived, protein-encoding circular RNAs are biologically functional and linked to the transforming properties of some HPV.

Access to Health Care, Beliefs, and Behaviors about Colorectal Cancer Screening among Korean Americans

Background: Colorectal cancer (CRC) is one of the most common cancers among Korean Americans (KAs) and their CRC screening rates are low. To raise the rates of CRC screening among KAs, it is necessary to improve our understanding of factors that influence their CRC screening behaviors. This study examined socio-demographics, access to health care, health and cultural beliefs, and behaviors about the fecal occult blood test (FOBT) for CRC screening among KAs aged 50 and older. Methods: Based on the health belief model, the cultural assessment model for health, and the Powe fatalism model, this study measured socio-demographics (age, gender, years in the U.S., marital status, education, employment, income, and acculturation), health care access (health insurance, having a regular doctor, physician recommendation, and trust in doctor), health beliefs (susceptibility, severity, benefits, barriers, and self-efficacy), and cultural beliefs (physical space, health temporal orientation, personal control, and fatalism) and FOBT. A cross-sectional survey (n=202) was conducted. Data analysis was conducted using descriptive analysis, Pearson correlation, and multivariate logistic regression. Results: This study found that physician recommendation was the strongest factor in lifetime FOBT utilization in KAs. The results also revealed a positive association among health temporal orientation, health fatalism, and lifetime FOBT among KAs, while previous research found a negative association between fatalism and cancer screening. Years in the U.S., employment, and having a regular doctor were significantly associated with having had a FOBT in the previous year. Conclusion: Study results suggested the need for public education programs to increase physician recommendation among KAs. Furthermore, the positive relationships among health temporal orientation, health fatalism, and FOBT utilization in KAs suggests that KAs have a desire to maintain health and find cancer early despite their fatalistic view on health.

Differential glucose requirement in skin homeostasis and injury identifies a therapeutic target for psoriasis.

Proliferating cells, compared with quiescent cells, are more dependent on glucose for their growth. Although glucose transport in keratinocytes is mediated largely by the Glut1 facilitative transporter, we found that keratinocyte-specific ablation of Glut1 did not compromise mouse skin development and homeostasis. Ex vivo metabolic profiling revealed altered sphingolipid, hexose, amino acid, and nucleotide metabolism in Glut1-deficient keratinocytes, thus suggesting metabolic adaptation. However, cultured Glut1-deficient keratinocytes displayed metabolic and oxidative stress and impaired proliferation. Similarly, Glut1 deficiency impaired in vivo keratinocyte proliferation and migration within wounded or UV-damaged mouse skin. Notably, both genetic and pharmacological Glut1 inactivation decreased hyperplasia in mouse models of psoriasis-like disease. Topical application of a Glut1 inhibitor also decreased inflammation in these models. Glut1 inhibition decreased the expression of pathology-associated genes in human psoriatic skin organoids. Thus, Glut1 is selectively required for injury- and inflammation-associated keratinocyte proliferation, and its inhibition offers a novel treatment strategy for psoriasis.

Cancer screening in Koreans: a focus group approach.

BACKGROUND: Cancer is the greatest disease burden in Korea. Cancer screening can reduce the burden of cancer but cancer screening rates among Koreans remain low. The purposes of this study were to a) understand Koreans' beliefs and knowledge about cancer screening, and b) explore preferred strategies for increasing cancer screening utilization. METHODS: We conducted a descriptive, qualitative study using eight face-to-face focus groups with a total of 64 Koreans aged 40 and over. Participants answered semi-structured, open-ended questions assessing their experiences with, and beliefs, knowledge, and opinions about, cancer screening. All interview data were recorded and analyzed in the context of the health belief model (HBM). RESULTS: The most important themes that emerged from the focus group data were (a) perceived susceptibility (most of the participants believed they were not susceptible to cancer; those who perceived themselves susceptible to cancer were reluctant to express it); (b) perceived benefits (early detection and feelings of relief after cancer screening were benefits; participants had screening because they wanted to take advantage of the Korean government's Medical Payment Support program for cancer patients who have participated in the National Cancer Screening program); (c) perceived barriers (no symptoms; self-care when having symptoms; widespread distrust of tests, doctors, and hospitals; unkind health care providers; the financial burdens of advanced cancer screening tests; and the discomfort during cancer screening); and (d) knowledge of the causes of cancer (incorrect knowledge including beliefs that stress, personality, and body overuse cause cancer). Almost all of the participants were very knowledgeable about the seriousness of cancer and were confident that they were able to have cancer screening. Participants preferred strategies of cancer screening using group interventions with family or friends; various information delivery methods; information emphasizing the importance of cancer prevention; convenient, free, or inexpensive services; and kind health care providers. CONCLUSIONS: This HBM-based research suggests that beliefs in low susceptibility to cancer, many barriers to cancer screening, and incorrect knowledge should be the foci for increasing cancer screening rates in Koreans. Interventions could change individual cultural beliefs and increase knowledge as well as the quality of health care for Koreans.

Glucose Uptake in Heterologous Expression Systems.

Understanding the physiological regulation of glucose transport requires the analysis of transporters, like GLUT1, in diverse tissue types. We document the utility of viral vectors for the stable expression of wild-type and modified GLUT1 transporter in different types of mammalian cells. Once expression of the alleles has been confirmed by Western blotting, the effect of specific mutations on the regulation of glucose transport can be measured through a previously described radiolabeled glucose transport assay. Although this discussion is focused on the GLUT1 transporter, these techniques are easily transferrable to other glucose transporters.

Instrument Adaptation, Modification, and Validation for Cultural Beliefs About Colorectal Cancer Screening Among Korean Americans.

BACKGROUND: Studies on colorectal cancer (CRC) screening among Korean Americans (KAs) lack culturally sensitive, reliable, and validated belief scales. OBJECTIVE: The purpose of this study was to adapt, modify, and validate instruments measuring cultural beliefs (physical space, health temporal orientation, personal control, colon cancer fatalism, and health fatalism) about CRC screening in KAs. METHODS: In phase I, instrument adaptation and modification (translation from English into Korean, individual interviews using cognitive interviewing, and expert reviews) were used to make existing cultural beliefs instruments culturally appropriate for KAs. In phase II, instrument validation (pilot test and cross-sectional survey) was used to examine the psychometric properties of the instrument among 202 KAs. RESULTS: Construct validity and reliability of the final Korean version of the instruments were examined using exploratory and confirmatory factor analyses and internal consistency reliability. Exploratory factor analysis using all of the cultural beliefs items resulted in 5 factors accounting for 46.55% of the variance. Factor loadings were greater than 0.40 for most items to be added to the scales reflecting Korean cultural perspectives. Cronbach's αs for all the cultural beliefs subscales were greater than .70. CONCLUSIONS: Findings from this study show that KAs have unique cultural beliefs that should be reflected in the instruments used for CRC screening research with this population. IMPLICATIONS FOR PRACTICE: The revised instrument could be useful in accurately measuring cultural beliefs among KAs and in developing culturally sensitive interventions to increase CRC screening behaviors among KAs.