Unique subcellular distribution of RPB1 with a phosphorylated C-terminal domain (CTD) in mouse oocytes during meiotic division and its relationship with chromosome separation.

Polymerase (RNA) II (DNA directed) polypeptide A (RPB1) is the largest subunit of RNA polymerase II (RNAPII), and phosphorylation of its C-terminal domain (CTD) is required for transcription initiation, elongation and RNA processing. Little is known about the CTD phosphorylation pattern and potential function during cell division when transcription is silenced. In this study, we assessed the protein expression and subcellular distribution of RPB1 during mouse oocyte meiotic division. Western blot analysis revealed that the RPB1 CTD was highly phosphorylated on Ser2 (pRPB1(Ser2)), Ser5 (pRPB1(Ser5)) and Ser7 (pRPB1(Ser7)). High and stable expression of pRPB1(Ser2) and pRPB1(Ser5) was detected from germinal vesicle (GV) to Metaphase II (MII) stage. In contrast, pRPB1(Ser7) only emerged after germinal vesicle breakdown (GVBD) and gradually increased to its peak level at metaphase I (MI) and MII. Immunofluorescence demonstrated that pRPB1(Ser2), pRPB1(Ser5) and pRPB1(Ser7) were pronouncedly aggregated within the nucleus of GV oocytes with a non-surrounded nucleolus (NSN) but very faintly labeled in oocytes with a surrounded nucleolus (SN). After meiotic resumption, pRPB1(Ser2) was again detected at spindle poles and co-localized with key microtubule organizing center (MTOC) components, pericentrin and γ-tubulin. pRPB1(Ser5) and pRPB1(Ser7) were assembled as filamentous aggregates and co-localized with microtubules throughout the spindle structure, responding to spindle-disturbing drugs, nocodazole or taxol, in pattern strongly similar to microtubules. pRPB1(Ser2) and pRPB1(Ser5) were constantly localized on chromosomes, with a relatively high concentration in centromere areas. Taken together, our data suggest that the CTD is highly phosphorylated and may be required for accurate chromosome segregation in mouse oocytes during meiosis.

Comparative studies of transformation behaviors and mechanisms of halophenols in multiple chemical oxidative systems.

Due to wide applications, halophenols (HPs), especially bromophenols, chlorophenols, and fluorophenols, are commonly detected but resistant to biological removal in wastewater treatment plants (WWTPs). This study investigated the overall transformation behaviors of three representative HPs (2,4-dichlorophenol: 24-DCP, 2,4-dibromophenol: 24-DBP, 2,4-difluorophenol: 24-DFP) in six chemical oxidative systems (KMnO4, K2FeO4, NaClO, O3, UV, and persulfate (PS)). The results revealed fast removal of selected HPs by O3, PS and K2FeO4, while a large discrepancy in their removal efficiencies occurred under UV irradiation, KMnO4 oxidation and particularly chlorination. Based on the analysis of the identified intermediates and products, coupling among the five routes was the general route, and dimers were the main intermediates for HP oxidation. The effect of the halogen atom on the transformation pathways of HPs was highly reaction type dependent. Among the six chemical treatments, PS could induce HPs to yield relatively low-molecular-weight polymers and obtain the highest coupling degree. Transition state (TS) calculations showed that the H atom linked to the phenoxy group of HPs was the most easily abstracted by hydroxyl radicals to form the coupling precursor, i.e., phenoxy radicals. This high coupling behavior further resulted in the increased toxicity to green algae. Characterization revealed that HP reaction solutions treated with PS had a severely negative effect on algae growth, photosynthetic pigment synthesis, and the antioxidant enzyme system. These findings can shed light on the reaction mechanisms of advanced oxidation technologies and some risk management and control of PS technique may be considered when treating phenolic pollutants.

CuS@TA-Fe Nanoparticle-Doped Multifunctional Hydrogel with Peroxide-Like Properties and Photothermal Properties for Synergistic Antimicrobial Repair of Infected Wounds.

Bacterial infection is a critical factor in wound healing. Due to the abuse of antibiotics, some pathogenic bacteria have developed resistance. Thus, there is an urgent need to develop a non-antibiotic-dependent multifunctional wound dressing for the treatment of bacteria-infected wounds. In this work, a multifunctional AOCuT hydrogel embedded with CuS@TA-Fe nanoparticles (NPs) through Schiff base reaction between gelatin quaternary ammonium salt - gallic acid (O-Gel-Ga) and sodium dialdehyde alginate (ADA) along with electrostatic interactions with CuS@TA-Fe NPs is prepared. These composite hydrogels possess favorable injectability, rapid shape adaptation, electrical conductivity, photothermal antimicrobial activity, and biocompatibility. Additionally, the doped NPs not only impart fast self-healing properties and excellent adhesion performance to the hydrogels, but also provide excellent peroxide-like properties, enabling them to scavenge free radicals and exhibit anti-inflammatory and antioxidant capabilities via photothermal (PTT) and photodynamic (PDT) effects. In an S. aureus infected wound model, the composite hydrogel effectively reduces the expression level of wound inflammatory factors and accelerates collagen deposition, epithelial tissue, and vascular regeneration, thereby promoting wound healing. This safe and synergistic therapeutic system holds great promise for clinical applications in the treatment of infectious wounds.

Acceleration of Oral Wound Healing under Diabetes Mellitus Conditions Using Bioadhesive Hydrogel.

Oral wounds under diabetic conditions display a significant delay during the healing process, mainly due to oxidative stress-induced inflammatory status and abnormal immune responses. Besides, the wet and complicated dynamic environment of the oral cavity impedes stable treatment of oral wounds. To overcome these, a biomimetic hydrogel adhesive was innovatively developed based on a mussel-inspired multifunctional structure. The adhesive displays efficient adhesion and mechanical harmony on the oral mucosa through enhanced bonding in an acidic proinflammatory environment. The bioadhesive hydrogel exhibits excellent antioxidative properties by mimicking antioxidative enzymatic activities to reverse reactive oxygen species (ROS)-mediated immune disorders. Experiments on oral wounds of diabetic rats showed that this hydrogel adhesive could effectively protect against mucosal wounds and obviously shorten the inflammatory phase, thus promoting the wound-healing process. Therefore, this study offers a promising therapeutic choice with the potential to advance the clinical treatment of diabetic oral wounds.

nHA-loaded gelatin/alginate hydrogel with combined physical and bioactive features for maxillofacial bone repair.

Critical-sized maxillofacial bone defects have been a tough clinical challenge considering their requirements for functional and structural repair. In this study, an injectable in-situ forming double cross-linked hydrogel was prepared from gelatin (Gel), 20 mg/mL alginate dialdehyde (ADA), 4.5 mg/mL Ca2+ and borax. Improved properties of composite hydrogel might well fit and cover irregular geometric shape of facial bone defects, support facial structures and conduct masticatory force. We innovatively constructed a bioactive poly-porous structure by decoration with nano-sized hydroxyapatite (nHA). The highly ordered, homogeneous and size-confined porous surface served as an interactive osteogenic platform for communication and interplay between macrophages and bone marrow derived stem cells (BMSCs). Effective macrophage-BMSC crosstalk well explained the remarkable efficiency of nHA-loaded gelatin/alginate hydrogel (nHA@Gel/ADA) in the repair of critical-size skull bone defect. Collectively, the composite hydrogel constructed here might serve as a promising alternative in repair process of complex maxillofacial bone defects.

Accurate and stable two-step LED position calibration method for Fourier ptychographic microscopy.

SIGNIFICANCE: Fourier ptychography microscopy (FPM) is a computational optical imaging technology that employs angularly varying illuminations and a phase retrieval algorithm to achieve a wide field of view and high-resolution imaging simultaneously. In the FPM, LED position error will reduce the quality of the reconstructed high-resolution image. To correct the LED positions, current methods consider each of the LED positions as independent and use an optimization algorithm to get each of the positions. When the positional misalignment is large or the search position falls into a local optimal value, the current methods may lack stability and accuracy. AIM: We improve the model of the LED position and propose an accurate and stable two-step correction scheme (tcFPM) to calibrate the LED position error. APPROACH: The improved LED positions model combines the overall offset, which represents the relative deviation of the LED array and the optical axis, with the slight deviation of each LED's independent position. In the tcFPM, the overall offset of the LED array is corrected at first, which obtains an approximate value of the overall offset of the LED array. Then the position of each LED is precisely adjusted, which obtains the slight offset of each LED. RESULTS: This LED position error model is more in line with the actual situation. The simulation and experimental results show that the method has high accuracy in correcting the LED position. Furthermore, the reconstruction process of tcFPM is more stable and significantly improves the quality of the reconstruction results, which is compared with some LED position error correction methods. CONCLUSIONS: An LED position error correction technology is proposed, which has a stable iterative process and improves the reconstruction accuracy of complex amplitude.

Biased activation of ß2-AR/Gi/GRK2 signal pathway attenuated ß1-AR sustained activation induced by ß1-adrenergic receptor autoantibody.

Heart failure is the terminal stage of many cardiac diseases, in which ß1-adrenoceptor (ß1-AR) autoantibody (ß1-AA) has a causative role. By continuously activating ß1-AR, ß1-AA can induce cytotoxicity, leading to cardiomyocyte apoptosis and heart dysfunction. However, the mechanism underlying the persistent activation of ß1-AR by ß1-AA is not fully understood. Receptor endocytosis has a critical role in terminating signals over time. ß2-adrenoceptor (ß2-AR) is involved in the regulation of ß1-AR signaling. This research aimed to clarify the mechanism of the ß1-AA-induced sustained activation of ß1-AR and explore the role of the ß2-AR/Gi-signaling pathway in this process. The beating frequency of neonatal rat cardiomyocytes, cyclic adenosine monophosphate content, and intracellular Ca2+ levels were examined to detect the activation of ß1-AA. Total internal reflection fluorescence microscopy was used to detect the endocytosis of ß1-AR. ICI118551 was used to assess ß2-AR/Gi function in ß1-AR sustained activation induced by ß1-AA in vitro and in vivo. Monoclonal ß1-AA derived from a mouse hybridoma could continuously activate ß1-AR. ß1-AA-restricted ß1-AR endocytosis, which was reversed by overexpressing the endocytosis scaffold protein ß-arrestin1/2, resulting in the cessation of ß1-AR signaling. ß2-AR could promote ß1-AR endocytosis, as demonstrated by overexpressing/interfering with ß2-AR in HL-1 cells, whereas ß1-AA inhibited the binding of ß2-AR to ß1-AR, as determined by surface plasmon resonance. ICI118551 biasedly activated the ß2-AR/Gi/G protein-coupled receptor kinase 2 (GRK2) pathway, leading to the arrest of limited endocytosis and continuous activation of ß1-AR by ß1-AA in vitro. In vivo, ICI118551 treatment attenuated myocardial fiber rupture and left ventricular dysfunction in ß1-AA-positive mice. This study showed that ß1-AA continuously activated ß1-AR by inhibiting receptor endocytosis. Biased activation of the ß2-AR/Gi/GRK2 signaling pathway could promote ß1-AR endocytosis restricted by ß1-AA, terminate signal transduction, and alleviate heart damage.

H3 Thr3 phosphorylation is crucial for meiotic resumption and anaphase onset in oocyte meiosis.

Haspin-catalyzed histone H3 threonine 3 (Thr3) phosphorylation facilitates chromosomal passenger complex (CPC) docking at centromeres, regulating indirectly chromosome behavior during somatic mitosis. It is not fully known about the expression and function of H3 with phosphorylated Thr3 (H3T3-P) during meiosis in oocytes. In this study, we investigated the expression and sub-cellular distribution of H3T3-P, as well as its function in mouse oocytes during meiotic division. Western blot analysis revealed that H3T3-P expression was only detected after germinal vesicle breakdown (GVBD), and gradually increased to peak level at metaphase I (MI), but sharply decreased at metaphase II (MII). Immunofluorescence showed H3T3-P was only brightly labeled on chromosomes after GVBD, with relatively high concentration across the whole chromosome axis from pro-metaphase I (pro-MI) to MI. Specially, H3T3-P distribution was exclusively limited to the local space between sister centromeres at MII stage. Haspin inhibitor, 5-iodotubercidin (5-ITu), dose- and time-dependently blocked H3T3-P expression in mouse oocytes. H3T3-P inhibition delayed the resumption of meiosis (GVBD) and chromatin condensation. Moreover, the loss of H3T3-P speeded up the meiotic transition to MII of pro-MI oocytes in spite of the presence of non-aligned chromosomes, even reversed MI-arrest induced with Nocodazole. The inhibition of H3T3-P expression distinguishably damaged MAD1 recruitment on centromeres, which indicates the spindle assembly checkpoint was impaired in function, logically explaining the premature onset of anaphase I. Therefore, Haspin-catalyzed histone H3 phosphorylation is essential for chromatin condensation and the following timely transition from meiosis I to meiosis II in mouse oocytes during meiotic division.