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Although the nucleolus is involved in ribosome biogenesis, the functions of numerous nucleolus-localized proteins remain unclear. In this study, we genetically isolated Arabidopsis thaliana salt hypersensitive mutant 1 (sahy1), which exhibits slow growth, short roots, pointed leaves, and sterility. SAHY1 encodes an uncharacterized protein that is predominantly expressed in root tips, early developing seeds, and mature pollen grains and is mainly restricted to the nucleolus. Dysfunction of SAHY1 primarily causes the accumulation of 32S, 18S-A3, and 27SB pre-rRNA intermediates. Coimmunoprecipitation experiments further revealed the interaction of SAHY1 with ribosome proteins and ribosome biogenesis factors. Moreover, sahy1 mutants are less sensitive to protein translation inhibitors and show altered expression of structural constituents of ribosomal genes and ribosome subunit profiles, reflecting the involvement of SAHY1 in ribosome composition and ribosome biogenesis. Analyses of ploidy, S-phase cell cycle progression, and auxin transport and signaling indicated the impairment of mitotic activity, translation of auxin transport carrier proteins, and expression of the auxin-responsive marker DR5::GFP in the root tips or embryos of sahy1 plants. Collectively, these data demonstrate that SAHY1, a nucleolar protein involved in ribosome biogenesis, plays critical roles in normal plant growth in association with auxin transport and signaling.

Wen-Shuo Kuo,1,2,* Tien-Sung Yeh,3,* Chia-Yuan Chang,4 Jui-Chang Liu,2 Chang-Hsin Chen,5 Edmund Cheung So,6,7 Ping-Ching Wu8 1School of Chemistry and Materials Science, Nanjing University of Information Science and Technology, Nanjing 210044, Jiangsu, People’s Republic of China; 2Allergy & Clinical Immunology Research Center, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan, Republic of China; 3Department of Physical Medicine and Rehabilitation, An Nan Hospital, China Medical University, Tainan 709, Taiwan, Republic of China; 4Department of Mechanical Engineering, National Cheng Kung University, Tainan 701, Taiwan, Republic of China; 5AbVision Inc., Milpitas, CA 95035, USA; 6Department of Anesthesia & Medicine Research, An Nan Hospital, China Medical University, Tainan 709, Taiwan, Republic of China; 7Graduate Institute of Medical Sciences, Chang Jung Christian University, Tainan 711, Taiwan, Republic of China; 8Department of Biomedical Engineering, National Cheng Kung University, Tainan, 701, Taiwan, Republic of China*These authors contributed equally to this workCorrespondence: Edmund Cheung So; Ping-Ching Wu Email edmundsotw@gmail.com; wbcxyz@bme.ncku.edu.twBackground: Although graphene quantum dots (GQDs) have received considerable research attention for their applications in various fields, the use of GQDs, such as nitrogen-doped GQDs (N-GQDs) and amino-functionalized N-GQDs (amino-N-GQDs), as photosensitizers to facilitate photodynamic therapy (PDT) has received limited research intention. To address this research gap, this study prepared novel amino-N-GQDs and investigated their properties.Methods: The amino-N-GQDs subjected to two-photon excitation (TPE) exhibited remarkable bactericidal capability in PDT. The bonding compositions of nitrogen and the amino-functionalized group played a critical role in their antimicrobial effects.Results: Compared with amino-group-free N-GQDs and amino-N-free GQDs, the amino-N-GQDs generated a higher amount of reactive oxygen species, demonstrating their superior efficacy for two-photon PDT. Additionally, the intrinsic luminescence properties and high photostability of the amino-N-GQDs demonstrate their suitability as an effective two-photon contrast agent for tracking bacteria during two-photon biomedical imaging.Conclusion: The amino-N-GQD and their remarkable properties may provide an efficient alternative approach for observing and easily eliminating malignant microbes in the future.Keywords: nitrogen atom-doped graphene quantum dots, amino-functionalized and nitrogen atom-doped graphene quantum dots, two-photon photodynamic therapy, two-photon contrast agent, two-photon biomedical imaging

Wen-Shuo Kuo, 1, 2 Chia-Yuan Chang, 3 Jui-Chang Liu, 4 Jian-Hua Chen, 5, 6,* Edmund Cheung So, 5– 7,* Ping-Ching Wu 8,* 1School of Chemistry and Materials Science, Nanjing University of Information Science and Technology, Nanjing, Jiangsu 210044, People’s Republic of China; 2Allergy & Clinical Immunology Research Center, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan Republic of China; 3Department of Mechanical Engineering, National Cheng Kung University, Tainan 701, Taiwan Republic of China; 4Department of Biochemistry and Molecular Biology, National Cheng Kung University, Tainan 701, Taiwan Republic of China; 5Department of Anesthesia & Medicine Research, An Nan Hospital, China Medical University, Tainan 709, Taiwan Republic of China; 6Department of Anesthesia, China Medical University, Taichung 404, Taiwan Republic of China; 7Graduate Institute of Medical Sciences, Chang Jung Christian University, Tainan 711, Taiwan Republic of China; 8Department of Biomedical Engineering, National Cheng Kung University, Tainan 701, Taiwan Republic of China*These authors contributed equally to this workCorrespondence: Jian-Hua Chen; Ping-Ching Wu Email aptx4869jfk@gmail.com; wbcxyz@bme.ncku.edu.twBackground: Multidrug-resistant (MDR) bacterial strain is a serious medical problem. Methicillin-resistant Staphylococcus aureus (MRSA) is resistant to many antibiotics and is often associated with several diseases such as arthritis, osteomyelitis, and endocarditis. The development of an alternative treatment for eliminating MDR bacteria such as MRSA has attracted a considerable amount of research attention. Moreover, the development of a material for highly efficient generation of reactive oxygen species (ROS) involving two-photon photodynamic therapy (PDT) is currently desirable.MaterialsandMethods: We present an example demonstrating that the use of water-soluble C 60(OH) 30 fullerenol with a 0.89 singlet oxygen quantum yield serving as a photosensitizer in PDT has the superior ability in effectively generating ROS.Results: It has ultra-low energy (228.80 nJ pixel− 1) and can perform 900 scans under two-photon excitation (TPE) in the near-infrared region (760 nm) to completely eliminate the MDR species. Furthermore, the favorable two-photon properties are absorption of approximately 760 nm in wavelength, absolute cross-section of approximately 1187.50 Göeppert–Mayer units, lifetime of 6.640 ns, ratio of radiative to nonradiative decay rates of approximately 0.053, and two-photon stability under TPE.Conclusion: This enabled water-soluble C 60(OH) 30 fullerenol to act as a promising two-photon photosensitizer proceeding with PDT to easily eliminate MDR species.Keywords: methicillin-resistant Staphylococcus aureus, two-photon photodynamic therapy, water-soluble fullerenol, two-photon excitation, near-infrared region

Silver nanoparticles (NPs) are installed into a porphyrinic zirconium-based metal–organic framework (Zr-MOF), NU-902, through a postsynthetic modification followed by an ion-exchange process. Both ...

Summary Background Baricitinib is an oral selective inhibitor of Janus kinase (JAK) 1 and JAK2, approved for the treatment of patients with active rheumatoid arthritis. Because baricitinib, like other disease-modifying antirheumatic drugs, is used chronically, continuous assessment of its long-term safety profile is important. Here we provide updated data supporting the existing safety profile of baricitinib in this patient population. Methods In this safety analysis, integrated data were included from nine phase 3, phase 2, and phase 1b clinical trials, and one long-term extension study with data up to 360 weeks, ending Feb 13, 2018. We analysed three integrated datasets, the largest of which was the all-bari-RA dataset, which includes patients who received any dose of baricitinib. We compared the safety of baricitinib with placebo on the basis of data from seven studies with baricitinib 4 mg and placebo and four studies with baricitinib 2 mg, including placebo to week 24 (placebo-controlled dataset). We did a dose-response assessment based on four studies with baricitinib 2 mg and 4 mg, including long-term extension data (2–4 mg extended dataset). We did an exploratory analysis of deaths and venous thromboembolic events in a subset of data from the all-bari-RA dataset that included patients who had ever taken baricitinib 2-mg or baricitinib 4-mg. We did an analysis for malignancies (excluding non-melanoma skin cancer) in the as-randomised population (patients not censored at rescue or dose change). Findings We collected data for 3770 patients who were given baricitinib for 10 127 patient-years of exposure in the all-bari-RA dataset (median 1115 days [IQR 426–1441], maximum 2520 days). The placebo-controlled dataset comprised 2836 patients, with 1215 in the placebo group, with 451 patient-years of exposure data; 479 in the baricitinib 2 mg group, with 186 patient-years of exposure data; and 1142 in the baricitinib 4 mg group, with 472 patient-years of exposure data. The 2–4 mg extended dataset comprised 958 patients, with 479 in both the 2 mg and 4 mg groups. No significant differences were seen for baricitinib 4 mg or 2 mg versus placebo, or for 4 mg versus 2 mg in the incidence of death, malignancy, serious infection, or major adverse cardiovascular events. Incidence of herpes zoster per 100 patient-years was higher for baricitinib (4 mg: 4·4 [95% CI 2·7–6·7]; 2 mg: 3·1 [1·1–6·8]) versus total placebo group (1·1 [0·4–2·5]), as were treatment-emergent infections (4 mg: 89·7 [81·3–98·6]; 2 mg: 84·0 [71·3–98·2] vs placebo 75·4 [67·6–83·9]). Consistent with previous reports, incidences in the all-bari-RA dataset for venous thromboembolic events was 0·5 (95% CI 0·4–0·6) per 100 patient-years, deep-vein thrombosis was 0·3 (0·2–0·5) per 100 patient-years, and pulmonary embolism was 0·2 (0·2–0·4) per 100 patient-years. Incidences of malignancy (excluding non-melanoma skin cancer) in the 2–4 mg extended dataset were 0·8 (0·4–1·5) per 100 patient-years for baricitinib 2 mg and 1·0 (0·5–1·7) per 100 patient-years for baricitinib 4 mg, without censoring patients who had dose changes or received rescue treatment. We found no indication of higher incidence of venous thromboembolic events in the baricitinib 4 mg group compared with the 2 mg group in the 2–4 mg extended dataset. Interpretation In this updated integrated analysis of patients with active rheumatoid arthritis exposed to baricitinib for a maximum of almost 7 years, baricitinib 2 mg and 4 mg maintained a similar safety profile to earlier analyses. No new safety signals were identified. Patients in the long-term extension study continue to be followed up to date. Funding Eli Lilly and Company, under license from Incyte Corporation.

The COVID-19 pandemic related measures had augmented the rise of online education. While online teaching has mitigated the negative impacts from educational institutional closures, it was unable to displace hands-on biomedical laboratory practical lessons effectively. Without practical sessions, there was concern over the imparting of laboratory skills even with video demonstrations. To investigate the effectiveness of different delivery modes in imparting laboratory skills, theoretical and practical student assessments were analyzed alongside an anonymous survey on their motivation and prior experience. The undergraduate students were exposed to 1) instructor-live demonstration, 2) video demonstration or 3) no demonstration prior to the practical test which was a plasmid extraction. Significantly higher mini-prep yields and purity were found for both instructor-live and video demonstrations compared to no demonstration. Comparison with pre-pandemic theoretical assessment performance showed no significant differences despite longer contact hours during pre-pandemic times. Prior lab experience and motivation for selecting the course did not significantly affect student mini-prep yields. In conclusion, our findings suggest that video demonstrations were as effective as instructor-live demonstrations during the pandemic without noticeably compromising the teaching and learning of biomedical laboratory skills.

Alopecia areata is an autoimmune condition characterized by rapid hair loss in the scalp, eyebrows, and eyelashes, for which treatments are limited. Baricitinib, an oral, selective, reversible inhibitor of Janus kinases 1 and 2, may interrupt cytokine signaling implicated in the pathogenesis of alopecia areata.We conducted two randomized, placebo-controlled, phase 3 trials (BRAVE-AA1 and BRAVE-AA2) involving adults with severe alopecia areata with a Severity of Alopecia Tool (SALT) score of 50 or higher (range, 0 [no scalp hair loss] to 100 [complete scalp hair loss]). Patients were randomly assigned in a 3:2:2 ratio to receive once-daily baricitinib at a dose of 4 mg, baricitinib at a dose of 2 mg, or placebo. The primary outcome was a SALT score of 20 or less at week 36.We enrolled 654 patients in the BRAVE-AA1 trial and 546 in the BRAVE-AA2 trial. The estimated percentage of patients with a SALT score of 20 or less at week 36 was 38.8% with 4-mg baricitinib, 22.8% with 2-mg baricitinib, and 6.2% with placebo in BRAVE-AA1 and 35.9%, 19.4%, and 3.3%, respectively, in BRAVE-AA2. In BRAVE-AA1, the difference between 4-mg baricitinib and placebo was 32.6 percentage points (95% confidence interval [CI], 25.6 to 39.5), and the difference between 2-mg baricitinib and placebo was 16.6 percentage points (95% CI, 9.5 to 23.8) (P0.001 for each dose vs. placebo). In BRAVE-AA2, the corresponding values were 32.6 percentage points (95% CI, 25.6 to 39.6) and 16.1 percentage points (95% CI, 9.1 to 23.2) (P0.001 for each dose vs. placebo). Secondary outcomes for baricitinib at a dose of 4 mg but not at a dose of 2 mg generally favored baricitinib over placebo. Acne, elevated levels of creatine kinase, and increased levels of low- and high-density lipoprotein cholesterol were more common with baricitinib than with placebo.In two phase 3 trials involving patients with severe alopecia areata, oral baricitinib was superior to placebo with respect to hair regrowth at 36 weeks. Longer trials are required to assess the efficacy and safety of baricitinib for alopecia areata. (Funded by Eli Lilly under license from Incyte; BRAVE-AA1 and BRAVE-AA2 ClinicalTrials.gov numbers, NCT03570749 and NCT03899259.).

Nitrogen doping and amino group functionalization through chemical modification lead to strong electron donation. Applying these processes to a large π-conjugated system of graphene quantum dot (GQD)-based materials as electron donors increases the charge transfer efficiency of nitrogen-doped amino acid-functionalized GQDs (amino-N-GQDs), resulting in enhanced two-photon absorption, post-two-photon excitation (TPE) stability, TPE cross-sections, and two-photon luminescence through the radiative pathway when the lifetime decreases and the quantum yield increases. Additionally, it leads to the generation of reactive oxygen species through two-photon photodynamic therapy (PDT). The sorted amino-N-GQDs prepared in this study exhibited excitation-wavelength-independent two-photon luminescence in the near-infrared region through TPE in the near-infrared-II region. The increase in size resulted in size-dependent photochemical and electrochemical efficacy, increased photoluminescence quantum yield, and efficient two-photon PDT. Therefore, the sorted amino-N-GQDs can be applicable as two-photon contrast probes to track and localize analytes in in-depth two-photon imaging executed in a biological environment along with two-photon PDT to eliminate infectious or multidrug-resistant microbes.

Additional file 4. Raw data of sub-fraction IE3-3G1 proteomics analysis using LC–MS/MS.

Background. Chronic kidney disease (CKD) is a critical public health issue with a huge financial burden for both patients and society worldwide. Unfortunately, there are currently no efficacious therapies to prevent or delay the progression of end-stage renal disease (ESRD). Traditional Chinese medicine practices have shown that Cordyceps militaris (C. militaris) mycelia have a variety of pharmacologically useful properties, including antitumor, immunomodulation, and hepatoprotection. However, the effect of mycelial C. militaris on CKD remains unclear. Methods. Here, we investigated the effects of C. militaris mycelia on mice with CKD using four types of media: HKS, HKS with vitamin A (HKS + A), CM, and CM with vitamin A (CM + A). Results. The results at day 10 revealed that the levels of blood urea nitrogen (BUN) were significantly lower in the HKS (41%), HKS + A (41%), and CM + A (34%) groups compared with those in the corresponding control groups (nephrectomic mice). The level of serum creatinine in the HKS + A group decreased by 35% at day 10, whereas the levels in the HKS and CM + A groups decreased only by 14% and 13%, respectively, on day 30. Taken together, this is the first report using four new media (HKS, HKS + A, CM, and CM + A medium) for C. militaris mycelia. Each medium of mycelial C. militaris on CKD exhibits specific effect on BUN, serum creatinine, body weight, total protein, and uric acid. Conclusions. Taken together, this is the first report using four new media (HKS, HKS + A, CM, and CM + A medium) for C. militaris mycelia. Each medium of mycelial C. militaris on CKD exhibits specific effects on BUN, serum creatinine, body weight, total protein, and uric acid. We concluded that treatment with C. militaris mycelia cultured in HKS or CM + A medium could potentially prevent the deterioration of kidney function in mice with CKD.

Additional file 2. Details of RNA-seq analysis.

Protein display, secretion and export in prokaryotes are essential for utilizing microbial systems as engineered living materials for medicines, biocatalysts, and protein factories. To select for improved signal peptides for Escherichia coli protein display, we utilized error-prone polymerase chain reaction (epPCR) coupled with single-cell sorting and microplate titer to generate, select, and detect improved Ag43 signal peptides. Through three rounds of mutagenesis and selection using green fluorescence from the 56 kDa sfGFP-beta-lactamase, we isolated clones that increased surface display from 1.4 to 3 folds as detected by the microplate plate-reader and native SDS-PAGE assays. To establish that the protein was displayed extracellularly, we trypsinised the bacterial cells to release the surface displayed proteins for analysis. This workflow demonstrated a fast and high-throughput method leveraging on epPCR and single-cell sorting to rapidly augment bacterial surface display, a method that could be applied to other bacterial proteins.

Additional file 1. Details of the identification of the anti-allergy fraction, IE3-3G1, from L. gasseri.

Aconiti kusnezoffii Radix (Caowu), a traditional Chinese medicine and Mongolian medicine, has been commonly used owing to its analgesic and anti-inflammatory effects. Here, we performed a qualitative analysis of aconite alkaloids (AAs) in raw and processed Aconiti kusnezoffii Radix by ultra-high-performance liquid chromatography (HPLC) electrospray ionization-tandem mass spectrometry. Furthermore, we quantified benzoylaconine, benzoylmesaconine, benzoylhypaconine, aconitine, mesaconitine, and hypaconitine in raw and processed Aconiti kusnezoffii Radix using a HPLC system coupled with a diode array detector. The analysis of AA fragmentation patterns in positive electrospray ionization mode revealed 155 AAs, including 6 with unambiguous identities, 109 with putative assignments, and 40 unknown compounds. Additionally, lipopolysaccharide-treated RAW264.7 macrophages were used to evaluate the anti-inflammatory properties of the alkaloid extracts of Aconiti kusnezoffii Radix. Importantly, alkaloid extracts of Aconiti kusnezoffii Radix at three different concentrations exhibited protective effect against lipopolysaccharide-induced inflammation. Thus, the proposed method can be used not only for the qualitative and quantitative analyses of Aconiti kusnezoffii Radix, but also for evaluating other Aconitum spp. Our study lays a foundation for the clinical application of poisonous Aconitum.

Additional file 3: Table S1. Experimental protocol of ion-exchange chromatography. Table S2. Experimental protocol of size-exclusion chromatography. Table S3. Data collection and refinement statistics of the LGp40 crystal. Figure S1. IL-12p40 levels of crude extracts used to stimulate mouse BMDC. Figure S2. Ion-exchange chromatography of crude extracts on a DEAE-Sepharose Fast Flow column. Figure S3. IL-12p40 levels of IE1-1 to IE4-2 fractions (fractions from ion-exchange chromatography) used to stimulate mouse BMDC. Figure S4. Size-exclusion chromatography of fractions from crude extract IE1 on a Sephacryl S-300 HR column. Figure S5. Size-exclusion chromatography of fractions from crude extract IE2 on a Sephacryl S-300 HR column. Figure S6. Size-exclusion chromatography of fractions from crude extract IE3 on a Sephacryl S-300 HR column. Figure S7. Identification and purification of sub-fraction IE3-3G1. Figure S8. GAPDH derived from probiotics and pathogens are dissimilar. Figure S9. RNA-seq analysis showed differentially regulated gene expression between LGp40 and CDp40-stimulated BMDM. Figure S10. The plasminogen interaction and plasmin activation ability of LGp40 decreased when the GAPDH activity was lost.

Hydrogen peroxide (H

To investigate the clinical effect of percutaneous endoscopic lumbar discectomy (PELD) through bone tunnel in the treatment of migrated lumbar intervertebral disc herniation.The clinical data of 42 patients with migrated lumbar intervertebral disc herniation treated through PELD techniques were retrospectively analyzed from October 2015 to December 2018. There were 26 males and 16 females, aged from 39 to 71 years old with a mean of(58.55±7.16) years. There were 7 cases where the affected segment was LThe operative time was from 69 to 105 min with a mean of (88.29±9.85) min;and no intraoperative complications such as neurovascular injury or dural tear were occurredin all patients. One case in the transpedicular approach group was changed to general anesthesia and translaminar approach due to local anesthesia intolerance. All the patients were followed up from 13 to 34 months, with a mean of (13.71±3.56) months. VAS and ODI were significantly improved in two groups immediately after surgery and 12 months after surgery (P0.05). According to modified Macnab criteria, 27 cases obtained excellent results, 11 good, 3 fair, and 1 poor. There were no postoperative complications such as lumbar fractures and postoperative infections in the follow-up patients.For migrated intervertebral disc herniation, the modified nucleus pulposus classification can be used to estimate the precise target before operation, and the reasonable bone tunnel approach can be selected to obtain good results.

BackgroundOptimising recombinant antibody production is important for cost-effective therapeutics and diagnostics. With impact on commercialisation, higher productivity beyond laboratory scales is highly sought, where efficient production can also accelerate antibody characterisations and investigations.MethodsInvestigating HEK293E cells for mammalian antibody production, various transfection and culture parameters were systematically analysed for antibody light chain production before evaluating them for whole antibody production. Transfection parameters investigated include seeding cell density, the concentration of the transfection reagent and DNA, complexation time, temperature, and volume, as well as culture parameters such as medium replacement, serum deprivation, use of cell maintenance antibiotic, incubation temperature, medium volume, post-transfection harvest day and common nutrient supplements.ResultsUsing 2 mL adherent HEK293E cell culture transfections with 25 kDa linear Polyethylenimine in the most optimised parameters, we demonstrated a ~2-fold production increase for light chain alone and for whole antibody production reaching 536 and 49 μg respectively in a cost-effective manner. With the addition of peptone, κ light chain increased by ~4-fold to 1032 μg while whole antibody increased to a lesser extent by ~2.5-fold to 51 μg, with benefits potentially for antibodies limited by their light chains in production.ConclusionsOur optimised findings show promise for a more efficient and convenient antibody production method through transfection and culture optimisations that can be incorporated to scale up processes and with potential transferability to other mammalian-based recombinant protein production using HEK293E cells.Statement of SignificanceRecombinant antibody production is crucial for antibody research and development. Systematically investigating transfection and culture parameters such as PEI/DNA concentrations, complexation time, volume, and temperature, supplements, etc., we demonstrated a ~4-fold light chain alone production increase to 1032 μg and a 2.5-fold whole antibody production increase to 51 μg from 2 mL transfections.

Chronic kidney disease (CKD) is a critical public health issue with a huge financial burden for both patients and society worldwide. Unfortunately, there are currently no efficacious therapies to prevent or delay the progression of end-stage renal disease (ESRD). Traditional Chinese medicine practices have shown thatHere, we investigated the effects ofThe results at day 10 revealed that the levels of blood urea nitrogen (BUN) were significantly lower in the HKS (41%), HKS + A (41%), and CM + A (34%) groups compared with those in the corresponding control groups (nephrectomic mice). The level of serum creatinine in the HKS + A group decreased by 35% at day 10, whereas the levels in the HKS and CM + A groups decreased only by 14% and 13%, respectively, on day 30. Taken together, this is the first report using four new media (HKS, HKS + A, CM, and CM + A medium) forTaken together, this is the first report using four new media (HKS, HKS + A, CM, and CM + A medium) for