Suprachoroidal and Subretinal Injections of AAV Using Transscleral Microneedles for Retinal Gene Delivery in Nonhuman Primates.

Retinal gene therapy using adeno-associated viruses (AAVs) is constrained by the mode of viral vector delivery. Intravitreal AAV injections are impeded by the internal limiting membrane barrier, while subretinal injections require invasive surgery and produce a limited region of therapeutic effect. In this study, we introduce a novel mode of ocular gene delivery in rhesus macaques using transscleral microneedles to inject AAV8 into the subretinal or suprachoroidal space, a potential space between the choroid and scleral wall of the eye. Using in vivo imaging, we found that suprachoroidal AAV8 produces diffuse, peripheral expression in retinal pigment epithelial (RPE) cells, but it elicited local infiltration of inflammatory cells. Transscleral subretinal injection of AAV8 using microneedles leads to focal gene expression with transduction of RPE and photoreceptors, and minimal intraocular inflammation. In comparison, intravitreal AAV8 shows minimal transduction of retinal cells, but elicits greater systemic humoral immune responses. Our study introduces a novel mode of transscleral viral delivery that can be performed without vitreoretinal surgery, with focal or diffuse transgene expression patterns suitable for different applications. The decoupling of local and systemic immune responses reveals important insights into the immunological consequences of AAV delivery to different ocular compartments surrounding the blood-retinal barrier

E-p-methoxycinnamic acid production in hairy root cultures of Scrophularia buergeriana Miquel

E-p-methoxycinnamic acid (MCA) is one of the main active constituents of Scrophularia buergeriana Miquel and has hepatoprotective, anti-amnestic, and neuroprotective activities. For studying in vitro production of MCA, we established a hairy root culture of S. buergeriana by infecting leaf explants with Agrobacterium rhizogenes R1000, and tested the growth and MCA production of these cultures using different strengths of media and concentrations of auxins. Hairy roots grown in half-strength MS medium showed the highest levels of growth (10.3 g/l) as well as MCA production (0.83 mg/100 g dry weight). Hairy root culture with the treatment of 0.5 mg/l IBA produced the highest amount of dry weight (11.8 g/l) and MCA (1.26 mg/100 g dry weight) production. These results demonstrate that the hairy root culture of S. buergeriana is a valuable alternative approach for the production of MCA.

Analysis of online parenting community posts on expanded newborn screening for metabolic disorders using topic modeling: a quantitative content analysis

Purpose As more newborns have received expanded newborn screening (NBS) for metabolic disorders, the overall number of false-positive results has increased. The purpose of this study was to explore the psychological impacts experienced by mothers related to the NBS process. Methods An online parenting community in Korea was selected, and questions regarding NBS were collected using web crawling for the period from October 2018 to August 2021. In total, 634 posts were analyzed. The collected unstructured text data were preprocessed, and keyword analysis, topic modeling, and visualization were performed. Results Of 1,057 words extracted from posts, the top keyword based on ‘term frequency-inverse document frequency’ values was “hypothyroidism,” followed by “discharge,” “close examination,” “thyroid-stimulating hormone levels,” and “jaundice.” The top keyword based on the simple frequency of appearance was “XXX hospital,” followed by “close examination,” “discharge,” “breastfeeding,” “hypothyroidism,” and “professor.” As a result of LDA topic modeling, posts related to inborn errors of metabolism (IEMs) were classified into four main themes: “confirmatory tests of IEMs,” “mother and newborn with thyroid function problems,” “retests of IEMs,” and “feeding related to IEMs.” Mothers experienced substantial frustration, stress, and anxiety when they received positive NBS results. Conclusion The online parenting community played an important role in acquiring and sharing information, as well as psychological support related to NBS in newborn mothers. Nurses can use this study’s findings to develop timely and evidence-based information for parents whose children receive positive NBS results to reduce the negative psychological impact

Seven-dimensional considerations of Einstein's connection. I. The recurrence relations of the first kind in n

Einstein's connection in a generalized Riemannian manifold Xn has been investigated by many authors for lower-dimensional cases n=2,…,6. In a series of papers, we obtain a surveyable tensorial representation of 7-dimensional Einstein's connection in terms of unified field tensor, with main emphasis on the derivation of powerful and useful recurrence relations which hold in X7. In this paper, we give a brief survey of Einstein's unified field theory and derive the recurrence relations of the first kind which hold in a general Xn. All considerations in this paper are restricted to the first and second classes only since the case of the third class, the simplest case, was already studied by many authors

NKT cells promote antibody-induced joint inflammation by suppressing transforming growth factor β1 production

Although NKT cells has been known to exert protective roles in the development of autoimmune diseases, the functional roles of NKT cells in the downstream events of antibody-induced joint inflammation remain unknown. Thus, we explored the functional roles of NKT cells in antibody-induced arthritis using the K/BxN serum transfer model. NKT cell–deficient mice were resistant to the development of arthritis, and wild-type mice administrated with α-galactosyl ceramide, a potent NKT cell activator, aggravated arthritis. In CD1d−/− mice, transforming growth factor (TGF)-β1 was found to be elevated in joint tissues, and the blockade of TGF-β1 using neutralizing monoclonal antibodies restored arthritis. The administration of recombinant TGF-β1 into C57BL/6 mice reduced joint inflammation. Moreover, the adoptive transfer of NKT cells into CD1d−/− mice restored arthritis and reduced TGF-β1 production. In vitro assay demonstrated that interleukin (IL)-4 and interferon (IFN)-γ were involved in suppressing TGF-β1 production in joint cells. The adoptive transfer of NKT cells from IL-4−/− or IFN-γ−/− mice did not reverse arthritis and TGF-β1 production in CD1d−/− mice. In conclusion, NKT cells producing IL-4 and IFN-γ play a role in immune complex–induced joint inflammation by regulating TGF-β1

Dietary Intake and Serum Levels of Iron in Relation to Oxidative Stress in Breast Cancer Patients

Iron may induce oxidative stress via production of reactive oxygen species, facilitating mammary carcinogenesis. This study investigated the role of iron in relation to oxidative stress as a potential risk factor in the development of breast cancer (BC). BC patients (n = 121) and healthy age-matched controls (n = 149) were entered into the study. Iron and antioxidant vitamins intakes were estimated using a quantitative food frequency questionnaire. Thirty one subjects from each group provided blood samples for measurement of serum iron, plasma malondialdehyde (MDA) and ferric reducing ability of plasma (FRAP). Total and non-heme iron intake of BC patients were lower than those of the controls. However, the serum iron level was significantly higher in BC patients. Plasma MDA levels were also significantly higher in BC patients whereas no significant difference in FRAP values were observed between the two groups. Log-transformed serum iron concentration showed no significant correlation with MDA or FRAP. These results suggest that serum iron overload may be a breast cancer risk factor possibly due to increased oxidative stress

Clinical Comparison of the Auditory Steady-State Response with the Click Auditory Brainstem Response in Infants

ObjectivesOur goal was to determine the effectiveness of using the auditory steady state response (ASSR) as a measure of hearing thresholds in infants who are suspected of having significant hearing loss, as compared with using the click-auditory brainstem response (C-ABR).MethodsWe retrospectively analyzed the audiologic profiles of 76 infants (46 boys and 30 girls, a total of 151 ears) who ranged in age from 1 to 12 months (average age: 5.7 months). The auditory evaluations in 76 infants who were suspected of having hearing loss were done via the C-ABR and ASSR. In addition, for reference, the mean ASSR thresholds were compared to those of 39 ears of infants and 39 ears of adults with normal hearing at 0.5, 1, 2, and 4 kHz.ResultsThe highest correlation between the C-ABR and ASSR thresholds was observed at an average of 2-4 kHz (r=0.94). On comparison between the hearing of infants and adults at 0.5, 1, 2, and 4 kHz, the mean ASSR threshold in infants was 12, 7, 8, and 7 dB higher, respectively, than that in adults.ConclusionASSR testing may provide additional audiometric information for accurately predicting the hearing sensitivity, and this is essential for the management of infants with severe to profound hearing loss

Crystal structures of murine norovirus-1 RNA-dependent RNA polymerase in complex with 2-thiouridine or ribavirin

AbstractMurine norovirus-1 (MNV-1) shares many features with human norovirus (HuNoV) and both are classified within the norovirus genus of Caliciviridae family. MNV-1 is used as the surrogate for HuNoV research since it is the only form that can be grown in cell culture. HuNoV and MNV-1 RNA dependent RNA polymerase (RdRp) proteins with the sequence identity of 59% show essentially identical conformations. Here we report the first structural evidence of 2-thiouridine (2TU) or ribavirin binding to MNV-1 RdRp, based on the crystal structures determined at 2.2Å and 2.5Å resolutions, respectively. Cellular and biochemical studies revealed stronger inhibitory effect of 2TU on the replication of MNV-1 in RAW 264.7 cells, compared to that of ribavirin. Our complex structures highlight the key interactions involved in recognition of the nucleoside analogs which block the active site of the viral RNA polymerase

Prevalence of human parechovirus and enterovirus in cerebrospinal fluid samples in children in Jinju, Korea

PurposeHuman parechovirus (HPeV) and enterovirus (EV) are causative agents of a sepsis-like illness in neonates and of infections of the central nervous system in young children. The objectives of this study were to assess the prevalence of HPeV3 and EV infection in young children with a sepsis-like illness or with meningitis in Jinju, Korea.MethodsCerebrospinal fluid (CSF) samples were collected from 267 patients (age range, 1 day to 5 years) and assessed for HPeV and EV by performing reverse transcription polymerase chain reaction assay. Amplification products of the VP3/VP1 region of HPeV and of the VP1 region of EV were sequenced to identify the virus type.ResultsHPeV and EV were detected in 3.4% and 7.5% of the total CSF samples assessed, respectively. The age distribution of EV-positive patients (median age, 1.4 months) had a significantly broader range than that of HPeV-positive patients (median age, 7.8 months). The peak seasons for HPeV and EV infection were spring and summer, respectively. The clinical symptoms for HPeV and EV infection were similar, and fever was the most common symptom. Pleocytosis was detected in 22.2% of HPeV-positive patients and 35.5% of EV-positive patients. The VP3/VP1 gene sequence of the nine Korean strains clustered most closely with the Japanese strain (AB759202).ConclusionThe data indicate that HPeV infection is predominant in young infants (<6 months) and that meningitis without pleocytosis was caused by both HPeV and EV infection in children