Autofluorescence microscopy as a non-invasive probe to characterize the complex mechanical properties of keratin-based integumentary organs: A feather paradigm.

Integumentary organs exhibit diverse morphologies and functions. The complex mechanical property of the architecture is mainly contributed by the ingenious multiscale assembly of keratins. A cross-scale characterization on keratin integration in an integument system will help us understand the principles on how keratin-based bio-architecture are built and function in nature. In this study, we used feather as a model integument organ. We develop autofluorescence (AF) microscopy to study the characteristics of its keratin assemblies over a wide range of length scales. The AF intensity of each feather component, following the hierarchy from the rachis to barb to barbule, decreased with the physical dimension. By combining the analysis of AF signal and tensile testing, we can probe regional material density and the associated mechanical strength in a composite feather. We further demonstrated that the AF micro-images could resolve subtle variations in the defective keratin assembly in feathers from frizzled chicken variants with a mutation in α-keratin 75. The distinction between AF patterns and the morphological features of feather components across different length scales indicated a synergetic interplay between material integration and complex morphogenesis during feather development. The work shows AF microscopy can serve as an easy and non-invasive approach to study multiscale keratin organizations and the associated bio-mechanical properties in diverse integumentary organs. This approach will facilitate our learning of many bio-inspired designs in diverse animal integumentary organs/appendages.

Novel human-derived EML4-ALK fusion cell lines identify ribonucleotide reductase RRM2 as a target of activated ALK in NSCLC.

INTRODUCTION: Echinoderm microtubule-associated protein-like 4 (EML4)-Anaplastic Lymphoma Kinase (ALK) rearrangements occur in 3% to 7% of lung adenocarcinomas and are targets for treatment with tyrosine kinase inhibitors (TKIs). Here we have developed three novel EML4-ALK-positive patient-derived Non-Small-Cell-Lung-Cancer (NSCLC) cancer cell lines, CUTO8 (variant 1), CUTO9 (variant 1) and CUTO29 (variant 3) and included a fourth ALK-positive cell line YU1077 (variant 3) to study ALK-positive signaling and responses. Variants 1 and 3 are the most common EML4-ALK variants expressed in ALK-positive NSCLC, and currently cell lines representing these EML4-ALK variants are limited. MATERIALS AND METHODS: Resazurin assay was performed to evaluate cell viability. Protein levels were determined using western blotting. RNA sequencing was performed in all four cell lines to identify differentially expressed genes. Whole-genome sequencing was performed to determine the presence of EML4-ALK fusion and ALK tyrosine kinase inhibitor resistance mutations. RESULTS: In this study, we have confirmed expression of the corresponding ALK fusion protein and assessed their sensitivity to a range of ALK tyrosine kinase inhibitors. These patient derived cell lines exhibit differential sensitivity to lorlatinib, brigatinib and alectinib, with EML4-ALK variant 3 containing cell lines exhibiting increased sensitivity to lorlatinib and brigatinib as compared to alectinib. These cell lines were further characterized by whole genome sequencing and RNA-seq analysis that identified the ribonucleotide reductase regulatory subunit 2 (RRM2) as a downstream and potential therapeutic target in ALK-positive NSCLC. CONCLUSION: We provide a characterization of four novel EML4-ALK-positive NSCLC cell lines, highlighting genomic heterogeneity and differential responses to ALK TKI treatment. The RNA-Seq characterization of ALK-positive NSCLC CUTO8, CUTO9, CUTO29 and YU1077 cell lines reported here, has been compiled in an interactive ShinyApp resource for public data exploration (https://ccgg.ugent.be/shiny/nsclc_rrm2_2022/).

Atomic replacement effects on the band structure of doped perovskite thin films.

The potential applications of perovskite manganite R1-xAxMnO3 (R = rare earth element; A = Sr, Ca) thin films have been continuously explored due to their multi-functional properties. In particular, the optimally hole-doped La0.67Ca0.33MnO3 thin film demonstrates a colossal magneto-resistance that is beneficial to the performance of spintronic devices. To understand the effect of R and A ions on the material properties, we systematically measure the resistivity, magnetization, and electronic energy states for three optimally hole-doped R0.67A0.33MnO3 thin films with R = La, Sm and A = Sr, Ca. Various energy parameters are derived based on the X-ray absorption and X-ray photoelectron spectra, including the band gap, the charge frustration energy and the magnetic exchange energy. It is interesting to find that the replacement of La with Sm is more effective than that of Sr with Ca in terms of tuning the electrical property, the Curie temperature, and the band gap. The strain-induced reduction of the O 2p- Mn 3d hybridization and the interplay of R/A site disorder and strain effect are discussed. The results of this study provide useful information for the band design of perovskite oxide films.

Activating natural killer (NK) cytotoxicity of canine CD5-CD21- cells requires low surface CD5 density NK cells.

Natural killer (NK) cells play a crucial role in regulating immune functions. Few studies have characterized canine NK cells. We previously demonstrated that canine peripheral blood lymphocytes (PBLs) with a low surface CD5 density (CD5lo) are considered a critical NK population. Natural killer cells in most mammals do not express T-cell markers, but canine CD5lo cells express surface molecules, such as CD3 T-cell receptors. These features make canines unique models for the study of comparative immunology in NK cells. In this study, we discovered that CD5lo and CD21 double-negative (CD5lo-ne/CD2-) cells were originally low in NK cytotoxicity and their NK cytotoxicity was highly activated when co-cultured with CD5lo NK cells. The cytotoxicity was not activated when co-cultured with other cell types, such as high surface CD5 density (CD5hi) cells. The CD5lo-negative (CD5lo-ne) population comprises CD5- and CD5hi cells. CD5-cells were low in NK cytotoxicity initially or after culturing with interleukin-2 (IL-2) without CD5lo cells; however, the addition of CD5lo cells in a similar medium markedly enhanced the NK activity. By contrast, CD5hi cells were always NK inactive, irrespective of them being cultured with CD5lo cells or not. We further verified that only the CD5-CD21- cells, which were separated from CD5-CD21+ cells in the entire CD5- population, showed activated NK activity through CD5lo cell induction. This study is the first to reveal that canine NK cells enhanced NK-inert cells to become NK-cytotoxic cells. Additionally, it is concluded that in beagles, except for CD5lo cells, CD5-CD21- cells show NK activity.

Association of dementia in patients with benign paroxysmal positional vertigo.

OBJECTIVE: We conducted a cohort study to investigate whether benign paroxysmal positional vertigo (BPPV) is correlated with an increased risk of dementia. METHODS: We established a case cohort comprising 7818 patients aged over 20 years who were diagnosed with BPPV from 2000 to 2010. In addition, we formed a control cohort by randomly selecting 31,272 people without BPPV and matched them with the BPPV patients according to gender, age, and index year. Cox proportional hazard regressions were performed to compute the hazard ratio (HR) of dementia after we adjusted for demographic characteristics and comorbidity. RESULTS: The prevalence of comorbidity was higher among patients with BPPV than among those without BPPV. In addition, patients with BPPV exhibited a 1.24-fold (95% confidence interval, CI 1.09-1.40; P < 0.001) higher risk of dementia than those without BPPV after we adjusted for age, gender, and comorbidity. An analysis stratified according to demographic factors revealed that women with BPPV exhibited a 1.36-fold (95% CI 1.16-1.59; P < 0.001) higher risk of dementia. Patients with BPPV aged over 65 years exhibited a significantly higher risk of dementia (adjusted HR: 1.26; 95% CI 1.10-1.43; P < 0.001) than those without BPPV. CONCLUSIONS: Patients with BPPV exhibited a higher risk of dementia than those without BPPV.

The Power of Serum Uric Acid in Predicting Metabolic Syndrome Diminishes With Age in an Elderly Chinese Population.

OBJECTIVES: Although serum uric acid (sUA) is not a criterion for diagnosing metabolic syndrome (MetS), many studies have identified a positive association between sUA and MetS in patients of various ages and ethnicities. This association has not been fully established in the very elderly. DESIGN: Cross-sectional and longitudinal study. SETTING AND PARTICIPANTS: A total of 18,906 Chinese elderly aged 65 and older undergoing routine health checkups in Taiwan were enrolled. MEASUREMENTS: Modified Adult Treatment Panel III criteria were used to define MetS. All participants were further divided into nine groups with gender specification according to age (the young-old, 65 to 74; old-old, 75 to 84; and oldest-old, 85 and over) and sUA concentration tertile (males: sUAG1, <5.7 mg/dL; sUAG2, 5.7-6.7 mg/dL; and sUAG3, > 6.7 mg/dL; females: sUAG1, <4.9 mg/dL; sUAG2, 4.9-5.9 mg/dL; and sUAG3, > 5.9 mg/dL). A cross-sectional study was first performed to determine the correlation between sUA and MetS and its components. A longitudinal study then excluded subjects with MetS at baseline to explore the risk of MetS according to sUA levels in 3 age groups. RESULTS: In the cross-sectional study, we observed a graded, positive association between sUA and MetS components that diminished after age 75. Subjects with higher sUA levels had higher odds ratios (OR) for the occurrence of MetS in the young-old and old-old groups of both sexes (P<0.001) except sUAG2 males in the old-old group. However, the association diminished with age and only a higher OR was observed in sUAG2 males in the oldest-old group (OR, 3.38; 95% CI, 1.11-10.30; P = 0.032). In the longitudinal study, the Kaplan-Meier plot showed that higher sUA levels were associated with a higher risk of MetS in the young-old group of both genders (P < 0.001 sUAG3 vs. sUAG1 and sUAG2). Cox regression analysis further confirmed these results (young-old group: sUAG3 HR, 1.90; 95% CI, 1.42-2.54; P < 0.001; old-old group males: HR, 2.20; 95% CI, 1.04-4.65; P = 0.039; young-old females: HR, 1.83; 95% CI, 1.38 - 2.43; P < 0.001). CONCLUSIONS: Higher sUA levels in the young-old group of Chinese elderly were associated with a higher risk of developing MetS. sUA levels are thus regarded as a potential tool for early diagnosis of MetS. However, this association diminished in those over 75 years of age.

Downregulation of COMMD1 by miR-205 promotes a positive feedback loop for amplifying inflammatory- and stemness-associated properties of cancer cells.

Sustained activation of nuclear factor-κB (NF-κB) in cancer cells has been shown to promote inflammation, expansion of cancer stem cell (CSC) population, and tumor development. In contrast, recent studies reveal that CSCs exhibit increased inflammation due to constitutive NF-κB activation; however, the underlying molecular mechanism remains unclear. In the present study, the analysis of microarray data revealed upregulation of NF-κB-regulated pro-inflammatory genes and downregulation of copper metabolism MURR1 domain-containing 1 (COMMD1) during the enrichment for stemness in SAS head and neck squamous-cell carcinoma (HNSCC) cells. The 3'-UTR of COMMD1 mRNA contains microRNA (miR)-205 target site. Parallel studies with HNSCC and NSCLC cells indicated that miR-205 is upregulated upon NF-κB activation and suppresses COMMD1 expression in stemness-enriched cancer cells. COMMD1 negatively regulates the inflammatory responses induced by TLR agonists, IL-1ß, and TNF-α by targeting RelA for degradation. The shRNA-mediated downregulation of COMMD1 in cancer cells enhanced inflammatory response, generating favorable conditions for macrophage recruitment. In addition, genes associated with stemness were also upregulated in these cells, which exhibited increased potential for anchorage-independent growth. Furthermore, COMMD1 downregulation promoted in vivo tumorigenesis and tumor growth, and tumors derived from COMMD1-knockdown cells displayed elevated level of NF-κB activation, increased expression of inflammatory- and stemness-associated genes, and contain expanded population of tumor-associated leukocytes and stemness-enriched cancer cells. These results suggest that COMMD1 downregulation by miR-205 promotes tumor development by modulating a positive feedback loop that amplifies inflammatory- and stemness-associated properties of cancer cells.

Long-term effects of maternal undernutrition on offspring carotid artery remodeling: role of miR-29c.

The purpose of this study was to examine the hypothesis that excess maternal glucocorticoids in response to maternal undernutrition programs the expression of extracellular matrix (ECM) components potentially by miR-29c. We measured the expression of mRNA (qRT-PCR) and protein (Western blot) for collagen 3A1, collagen 4A5 and matrix metalloproteinase 2 (MMP2) in offspring carotid arteries from three groups of dams: 50% food-restricted in latter half of gestation [maternal undernutrition (MUN)], MUN dams who received metyrapone (MET) (500 mg/ml ) in drinking water from day 10 of gestation to term, and control dams fed an ad libitum diet. The expression of miR-29c was significantly decreased at 3 weeks, 3 months and 9 months in MUN carotid arteries, and these decreases in expression were partially blocked by treatment of dams with MET. The expression pattern of ECM genes that are targets of miR-29c correlated with miR-29c expression. Expression of mRNA was increased for elastin (ELN) and MMP2 mRNA in 3-week MUN carotids; in 9-month carotids there were also significant increases in expression of Col3A1 and Col4A5. These changes in mRNA expression of ECM genes at 3 weeks and 9 months were blocked by MET treatment. Similarly, the expression of ELN and MMP2 proteins at 3 weeks were increased in MUN carotids, and by 9 months there were also increases in expression of Col3A1 and Col4A5, which were blocked by MET in MUN carotids. Overall, the results demonstrate a close correlation between expression of miR-29c and the ECM proteins that are its targets thus supporting our central hypothesis.

Long-living terahertz magnons in ultrathin metallic ferromagnets.

The main idea behind magnonics is to use the elementary magnetic excitations (magnons) for information transfer and processing. One of the main challenges, hindering the application of ultrafast terahertz magnons in magnonics, has been the short lifetime of these excitations in metallic ferromagnets. Here, we demonstrate that the engineering of the electronic structure of a ferromagnetic metal, by reducing its dimensionality and changing its chemical composition, opens a possibility to strongly suppress the relaxation channels of terahertz magnons and thereby enhance the magnons' lifetime. For the first time, we report on the long-lived terahertz magnons excited in ultrathin metallic alloy films. On the basis of the first-principles calculations, we explain the microscopic nature of the long lifetime being a consequence of the peculiar electronic hybridizations of the species. We further demonstrate a way of tailoring magnon energies (frequencies) by varying the chemical composition of the film.

Evaluation of salivary function by sialoscintigraphy in locally advanced nasopharyngeal cancer patients after intensity modulated radiotherapy.

PURPOSE: This study aimed to evaluate the salivary gland function changes by sialoscintigraphy in locally advanced nasopharyngeal cancer (NPC) after intensity modulated radiotherapy (IMRT). MATERIALS AND METHODS: Salivary function was assessed by sialoscintigraphy. Quantitative sialoscintigraphy was performed in 24 NPC patients prior to and after IMRT. Results were categorized in four groups according to the duration of treatment. The sialoscintigraphy parameters were examined. RESULTS: Sialoscintigraphy showed a significant difference in the secretion of each interval groups. The parameters of scintigraphy, except maximum accumulation (MA) of submandibular glands, decreased first after radiotherapy, and then recovered. However, the MA of submandibular glands was continuously downhill after radiation. CONCLUSIONS: The sialoscintigraphy parameters of each gland changed with the different radiation dose and follow-up intervals. The salivary function was influenced after radiotherapy in locally advanced NPC, especially, in the submandibular gland. Strategies to improve the salivary function should be assessed.

MicroRNA-19a-3p inhibits breast cancer progression and metastasis by inducing macrophage polarization through downregulated expression of Fra-1 proto-oncogene.

One of the hallmarks of malignancy is the polarization of tumor-associated macrophages (TAMs) from a pro-immune (M1-like) phenotype to an immune-suppressive (M2-like) phenotype. However, the molecular basis of the process is still unclear. MicroRNA (miRNA) comprises a group of small, non-coding RNAs that are broadly expressed by a variety of organisms and are involved in cell behaviors such as suppression or promotion of tumorigenesis. Here, we demonstrate that miR-19a-3p, broadly conserved among vertebrates, was downregulated in RAW264.7 macrophage cells of the M2 phenotype in conditoned medium of 4T1 mouse breast tumor cells. This downregulation correlated with an increased expression of the Fra-1 gene, which was reported to act as a pro-oncogene by supporting the invasion and progression of breast tumors. We found significant upregulation of miR-19a-3p in RAW264.7 macrophages after transfection with a miR-19a-3p mimic that resulted in a significant suppression of the expression of this gene. In addition, we could measure the activity of binding between miR-19a-3p and Fra-1 with a psiCHECK luciferase reporter system. Further, transfection of RAW264.7 macrophage cells with the miR-19a-3p mimic decreased the expression of the Fra-1 downstream genes VEGF, STAT3 and pSTAT3. Most importantly, the capacity of 4T1 breast tumor cells to migrate and invade was impaired in vivo by the intratumoral injection of miR-19a-3p. Taken together, these findings indicate that miR-19a-3p is capable of downregulating the M2 phenotype in M2 macrophages and that the low expression of this miRNA has an important role in the upregulation of Fra-1 expression and induction of M2 macrophage polarization.

Direct probing of the exchange interaction at buried interfaces.

The fundamental interactions between magnetic moments at interfaces have an important impact on the properties of layered magnetic structures. Hence, a direct probing of these interactions is highly desirable for understanding a wide range of phenomena in low-dimensional solids. Here we propose a method for probing the magnetic exchange interaction at buried interfaces using spin-polarized electrons and taking advantage of the collective nature of elementary magnetic excitations (magnons). We demonstrate that, for the case of weak coupling at the interface, the low-energy magnon mode is mainly localized at the interface. Because this mode has the longest lifetime of the modes and has a finite spectral weight across the layers on top, it can be probed by electrons. A comparison of experimental data and first-principles calculations leads to the determination of the interface exchange parameters. This method may help the development of spectroscopy of buried magnetic interfaces.

Neuroprotective effect of agmatine in rats with transient cerebral ischemia using MR imaging and histopathologic evaluation.

PURPOSE: This study aimed to further investigate the effects of agmatine on brain edema in the rats with middle cerebral artery occlusion (MCAO) injury using magnetic resonance imaging (MRI) monitoring and biochemical and histopathologic evaluation. MATERIALS AND METHODS: Following surgical induction of MCAO for 90min, agmatine was injected 5min after beginning of reperfusion and again once daily for the next 3 post-operative days. The events during ischemia and reperfusion were investigated by T2-weighted images (T2WI), serial diffusion-weighted images (DWI), calculated apparent diffusion coefficient (ADC) maps and contrast-enhanced T1-weighted images (CE-T1WI) during 3h-72h in a 1.5T Siemens MAGNETON Avanto Scanner. Lesion volumes were analyzed in a blinded and randomized manner. Triphenyltetrazolium chloride (TTC), Nissl, and Evans Blue stainings were performed at the corresponding sections. RESULTS: Increased lesion volumes derived from T2WI, DWI, ADC, CE-T1WI, and TTC all were noted at 3h and peaked at 24h-48h after MCAO injury. TTC-derived infarct volumes were not significantly different from the T2WI, DWI-, and CE-T1WI-derived lesion volumes at the last imaging time (72h) point except for significantly smaller ADC lesions in the MCAO model (P<0.05). Volumetric calculation based on TTC-derived infarct also correlated significantly stronger to volumetric calculation based on last imaging time point derived on T2WI, DWI or CE-T1WI than ADC (P<0.05). At the last imaging time point, a significant increase in Evans Blue extravasation and a significant decrease in Nissl-positive cells numbers were noted in the vehicle-treated MCAO injured animals. The lesion volumes derived from T2WI, DWI, CE-T1WI, and Evans blue extravasation as well as the reduced numbers of Nissl-positive cells were all significantly attenuated in the agmatine-treated rats compared with the control ischemia rats (P<0.05). CONCLUSION: Our results suggest that agmatine has neuroprotective effects against brain edema on a reperfusion model after transient cerebral ischemia.

Impact of atomic structure on the magnon dispersion relation: a comparison between Fe(111)/Au/W(110) and Fe(110)/W(110).

We present a combined experimental and theoretical study of the interplay between the atomic structure and the magnon excitations in low dimensional ferromagnets. Two monolayer thick Fe films on W(110) with and without a Au buffer layer are investigated. Our experiments show that adding the Au layer leads to a significant softening of the magnons. First-principles calculations confirm the experimental results revealing a strong dependency of exchange interactions on the atomic structure. It is observed that the intralayer exchange interactions increase with increasing distance between Fe layers. This unusual relationship is attributed to the complexity of the electronic structure and the contribution of different orbitals to the hybridization and exchange interaction. Our results suggest a way of tailoring magnetic excitations in low-dimensional magnetic structures.

Relaxation time of terahertz magnons excited at ferromagnetic surfaces.

The temporal and spatial properties of terahertz magnons excited at ferromagnetic fcc Co(100) and bcc Fe(110) surfaces are investigated experimentally. The magnon lifetime is found to be a few tens of femtoseconds at low wave vectors, which reduces significantly as the wave vector approaches the Brillouin zone boundary. Surprisingly, the lifetime is very similar in both systems, in spite of the fact that the excitation energy in the Co(100) film is by a factor of two larger than in the Fe(110) film. The magnon wave packets propagate only a few nanometers within their lifetime. In addition to the fact that our results describe the damping mechanism in ultrafast time scales, they may provide a way to predict the ultimate time scale of magnetic switching in nanostructures.

Magnon lifetimes on the Fe(110) surface: the role of spin-orbit coupling.

We provide direct experimental evidence which demonstrates that, in the presence of a large spin-orbit coupling, the lifetime, amplitude, group, and phase velocity of the magnons propagating along two opposite (but crystallographically equivalent) directions perpendicular to the magnetization are different. A real time and space representation reveals that magnons with the same energy (eigenfrequency) propagate differently along two opposite directions. Our findings can inspire ideas for designing new spintronic devices.