Complex interplay of neurodevelopmental disorders (NDDs), fractures, and osteoporosis: a mendelian randomization study.

BACKGROUND: Neurodevelopmental disorders (NDDs), such as Attention-Deficit/Hyperactivity Disorder (ADHD), Autism Spectrum Disorder (ASD), and Tourette Syndrome (TS), have been extensively studied for their multifaceted impacts on social and emotional well-being. Recently, there has been growing interest in their potential relationship with fracture risks in adulthood. This study aims to explore the associations between these disorders and fracture rates, in order to facilitate better prevention and treatment. METHODS: Employing a novel approach, this study utilized Mendelian randomization (MR) analysis to investigate the complex interplay between ADHD, ASD, TS, and fractures. The MR framework, leveraging extensive genomic datasets, facilitated a systematic examination of potential causal relationships and genetic predispositions. RESULTS: The findings unveil intriguing bidirectional causal links between ADHD, ASD, and specific types of fractures. Notably, ADHD is identified as a risk factor for fractures, with pronounced associations in various anatomical regions, including the skull, trunk, and lower limbs. Conversely, individuals with specific fractures, notably those affecting the femur and lumbar spine, exhibit an increased genetic predisposition to ADHD and ASD. In this research, no correlation was found between TS and fractures, or osteoporosis.These results provide a genetic perspective on the complex relationships between NDDs and fractures, emphasizing the importance of early diagnosis, intervention, and a holistic approach to healthcare. CONCLUSION: This research sheds new light on the intricate connections between NDDs and fractures, offering valuable insights into potential risk factors and causal links. The bidirectional causal relationships between ADHD, ASD, and specific fractures highlight the need for comprehensive clinical approaches that consider both NDDs and physical well-being.

Neurodevelopmental disorders as a risk factor for temporomandibular disorder: evidence from Mendelian randomization studies.

Objective: This study aims to clarify the incidence rate of temporomandibular joint disease in patients with mental disorders. Methods: Data extracted from the Psychiatric Genomics Consortium and FinnGen databases employed the Mendelian Randomization (MR) method to assess the associations of three neurodevelopmental disorders (NDDs)-Attention-Deficit/Hyperactivity Disorder (ADHD), Autism Spectrum Disorder (ASD), and Tourette's Disorder (TD)-as exposure factors with Temporomandibular Disorder (TMD). The analysis used a two-sample MR design, employing the Inverse Variance Weighted (IVW) method to evaluate the relationships between these disorders and Temporomandibular Disorder. Sensitivity analysis and heterogeneity assessments were conducted. Potential confounding factors like low birth weight, childhood obesity, and body mass index were controlled for. Results: The study found that ADHD significantly increased the risks for TMD (OR = 1.2342, 95%CI (1.1448-1.3307), p < 0.00001), TMD (including avohilmo) (OR = 1.1244, 95%CI (1.0643-1.1880), p = 0.00003), TMD-related pain (OR = 1.1590, 95%CI (1.0964-1.2252), p < 0.00001), and TMD-related muscular pain associated with fibromyalgia (OR = 1.1815, 95%CI (1.1133-1.2538), p < 0.00001), while other disorders did not show significant causal relationships. Conclusion: This study reveals the elevated risk of various TMD aspects due to ADHD. Furthermore, we discuss the link between low vitamin D levels ADHD and TMD. Future research should address these limitations and delve further into the complex interactions between ADHD, ASD, TD, and TMD.

Unveiling the muscle-brain axis: A bidirectional mendelian randomization study investigating the causal relationship between sarcopenia-related traits and brain aging.

BACKGROUND: Observational studies suggest an association between sarcopenia-related traits and brain aging, but whether this association reflects a causal relationship remains unclear. This study aims to employ Mendelian randomization (MR) methods to investigate the causal impact of sarcopenia-related traits on brain aging. METHODS: This study presents a comprehensive analysis of genome-wide association study (GWAS) summary data associated with sarcopenia-related traits. The data were derived from a large-scale cohort, encompassing measures such as grip strength, lean body mass, and walking pace. Measurements of brain aging were obtained from neuroimaging genetics, utilizing meta-analysis (ENIGMA) to combine magnetic resonance imaging (MRI) data from 33,992 participants. The primary methodology employed in this analysis was the inverse-variance-weighted method (IVW). Additionally, sensitivity analyses were conducted, to assess heterogeneity and pleiotropy. RESULT: Appendicular lean mass(ALM) is negatively correlated with Pallidum aging; Whole body fat-free mass shows a negative correlation with Amygdala aging; Leg fat-free mass (left) and Leg fat-free mass (right) are negatively correlated with Pallidum aging; Usual walking pace is positively correlated with Nucleus Accumbens aging. Cerebellum WM aging is negatively correlated with Leg fat-free mass (left) and Leg fat-free mass (right); Hippocampus aging is negatively correlated with Hand grip strength (left) and Hand grip strength (right). Ventricles aging is positively correlated with Usual walking pace; Nucleus Accumbens aging is positively correlated with Leg fat-free mass (left) and Leg fat-free mass (right); Putamen aging is positively correlated with ALM. CONCLUSION: Our study confirms that reduced muscle mass speeds up brain aging. Walking too fast raises the risk of brain aging, while maintaining or increasing appendicular lean mass, overall muscle mass, and muscle mass in both legs lowers the risk of brain aging.

USP10 regulates macrophage inflammation responses via stabilizing NEMO in LPS-induced sepsis.

BACKGROUND: Sepsis is a systemic inflammatory response syndrome characterized by persistent inflammation and immunosuppression, leading to septic shock and multiple organ dysfunctions. Ubiquitin-specific peptidase 10 (USP10), a deubiquitinase enzyme, plays a vital role in cancer and arterial restenosis, but its involvement in sepsis is unknown. OBJECTIVE: In this study, we investigated the significance of USP10 in lipopolysaccharide (LPS)-stimulated macrophages and its biological roles in LPS-induced sepsis. METHODS: Lipopolysaccharides (LPS) were used to establish sepsis models in vivo and in vitro. We use western blot to identify USP10 expression in macrophages. Spautin-1 and USP10-siRNA were utilized for USP10 inhibition. ELISA assays were used to assess for TNF-α and IL-6 in vitro and in vivo. Nuclear and cytoplasmic protein extraction and Confocal microscopy were applied to verify the translocation of NF-κB. Mechanically, co-immunoprecipitation and rescue experiments were used to validate the regulation of USP10 and NEMO. RESULTS: In macrophages, we found that LPS induced USP10 upregulation. The inhibition or knockdown of USP10 reduced the pro-inflammatory cytokines TNF-α and IL-6 and suppressed LPS-induced NF-κB activation by regulating the translocation of NF-κB. Furthermore, we found that NEMO, the regulatory subunit NF-κB essential modulator, was essential for the regulation of LPS-induced inflammation by USP10 in macrophages. NEMO protein evidently interacted with USP10, whereby USP10 inhibition accelerated the degradation of NEMO. Suppressing USP10 significantly attenuated inflammatory responses and improved the survival rate in LPS-induced sepsis mice. CONCLUSIONS: Overall, USP10 was shown to regulate inflammatory responses by stabilizing the NEMO protein, which may be a potential therapeutic target for sepsis-induced lung injury.

Integrated UPLC-MS, network pharmacology, and intestinal flora analysis to determine the treatment effect of Qiangzhi decoction on comorbid Tourette syndrome and RRTI.

Background: Tourette syndrome (TS) is a complex neurodevelopmental disorder characterized by vocal and motor tics. Recurrent respiratory tract infection (RRTI), a commonly occurring disease in childhood, correlates with recurrent and severe course of tic symptoms. Qiangzhi decoction (QZD) is a traditional Chinese medicine that can alleviate TS symptoms while reducing the recurrence of RRTI. However, the mechanism of QZD on TS and RRTI remains unclear. This study aimed to determine the treatment effect of QZD on comorbid TS and RRTI by integrating ultrahigh-performance liquid chromatography mass spectrometry (UPLC-MS), network pharmacology, and intestinal flora analysis. Methods: The components of QZD were first identified by UPLC-quadrupole (Q)-orbitrap-MS/MS. The mechanism of QZD on comorbid RRTI and TS was investigated by a series of network pharmacological methods, including target prediction and bioinformatics analysis. Finally, a comorbid TS and RRTI rat model was established by intraperitoneal injection of 3,3-iminodipropionitrile (IDPN), cyclophosphamide (CTX), and lipopolysaccharide (LPS). Alteration of gut microbiota in the alleviation of TS and RRTI by QZD was investigated via intestinal flora analysis. Results: The results of UPLC-Q-orbitrap-MS/MS showed that QZD had 96 types of chemical components. The network pharmacology results demonstrated that targets of QZD involved in the treatment of TS and RRTI involved 1045 biological processes (BPs), 109 cellular components (CCs), and 133 molecular functions (MFs), including synaptic and transsynaptic signaling, chemical synaptic transmission, neurotransmitter receptor activity, G protein-coupled amine receptor activity, and serotonin receptor activity, among others. Firmicutes, Bacteroidetes, Coprococcus, and Lachnospiraceae played crucial roles in gut microbiota of a QZD-treated comorbid TS and RRTI model. Conclusions: Our results revealed QZD provided a multicomponent, multitarget, and multipathway synergistic treatment of comorbid TS and RRTI.

Bioactivities and mechanism of action of securinega alkaloids derivatives reported prior to 2022.

Securinega alkaloids are indolizidine alkaloids extracted from the leaf and root of an Asian plant, Securinega suffruticosa. Since its discovery in 1956 by Russian scientists, numerous studies have been conducted on securinega alkaloids and their derivatives as bioactive agents. In this review, published work on the bioactivities and the mechanism of action of securinega alkaloids and their derivatives is addressed. References were obtained through for example, the Web of Science, Science Direct, Pubmed and Google Scholar. Research into the synthesis of securinega alkaloids and their derivatives lacking activity assessment has been excluded. Comprehensive reviews show that securinega alkaloids and their derivatives exhibit a wide range of activities among which antineoplastic activity and nervous system related activity were reported although the mechanisms of action remain in part unknown. The other activities such as induction of differentiation, reversal of multi-drug resistance, cardiovascular system related activity, anti-inflammatory, adjuvant agent and anti-pathogenic activity are also reviewed. We found that modification at the C12, C14, and C15 sites on securinine improves the antitumor activity, while derivatives in which a bivalent mimetic is linked to the C15 site is beneficial for differentiation induction activity and reversal of P-glycoprotein mediated drug resistance. The most related pathways involved in the bioactivity of securinega alkaloids and their derivatives are JAK/STAT, PI3K/AKT/mTOR and MAPK. A perspective and expectation concerning the research of securinega alkaloids is presented at the end of this article. This review indicates directions around which constant endeavor could be valuable for researchers in the near future.

Non-linear association between composite dietary antioxidant index and depression.

Background: Growing evidence has shown that the antioxidant diet is a protective factor against depression. However, the relationship between the Composite Dietary Antioxidant Index (CDAI), an important measure of antioxidant diet, and depression has received little attention. Therefore, we investigated the relationship between CDAI and depression through a cross-sectional analysis of the National Health and Nutrition Examination Survey (NHANES) from 2007 to 2018. Methods: The association between CDAI and depression was investigated using a weighted multiple logistic regression model with subgroup analysis. Non-linear correlations were explored using fitted smoothing curves. And we used a recursive method to figure out the turning point and build a weighted two-piece linear regression model. Results: In the multivariate logistic regression model with full adjustment for confounding variables, the ORs (95% CI) for the association between CDAI and depression were 0.83 (0.78, 0.88). Moreover, a non-linear association was found, with 0.16 being the inflection point. Before the inflection point, each unit increase in CDAI was associated with a 30% decrease in the risk of depression. After the inflection point, the risk of depression was found to be reduced by 11% for each unit increase. None of the interactions in all subgroup analyses were statistically significant. Conclusions: Our study highlighted a negative non-linear association between CDAI and depression in a nationally representative sample of US adults. Further clinical and basic research is needed to explore their association better.

Recombinant jurkat cells (HMGN2-T cells) secrete cytokines and inhibit the growth of tumor cells.

High Mobility Group Chromosomal Protein N2 (HMGN2) can recognize tumor cells and enhance the anti-tumor effect of immune cells. This study aimed to establish a lentiviral vector of recombinant HMGN2 gene, establish recombinant T cells (HMGN2-T cells), and observe their anti-tumor effects. Total RNA was isolated from peripheral blood mononuclear cells. HMGN2, cluster of differentiation (CD) 8 A, CD28, CD137, and CD3ζ genes were amplified and connected. Jurkat cells were transfected with the recombinant lentivirus vector. The viability, apoptosis, and cell cycle of HMGN2-T cells were detected using Cell Counting Kit-8 assay and flow cytometry. The co-culture was performed by adding HMGN2-T cells to tumor cells with different effect-to-target (E:T) ratios. The cytotoxic activity was measured by lactate dehydrogenase (LDH) releasing assay. The sequences of HMGN2, CD8A, CD28, CD137, and CD3ζ gene plasmids were confirmed using gene sequencing. After the lentiviral transfection for 72 h, green fluorescence cells (HMGN2-T cells) could be seen. Cell viability and apoptosis were increased in HMGN2-T cells. The cytokine levels of interleukin 2 (IL-2) and tumor necrosis factor α (TNF-α) increased in cell supernatants of HMGN2-T cells. The percentage of G0/G1 phase cells was lower, the rate of S phase cells was higher in HMGN2-T cells than control cells. The co-culture of HMGN2-T cells and tumor cells could promote the cytokines' release. The LDH level was increased with the elevation of E:T ratios. In conclusion, the HMGN2-T cells were well-established and have the effect of secreting cytokines and killing tumor cells.

A Super-Microsurgery Training Model: The Mouse Caudal Artery Anastomosis Model.

Objective: To establish a simple and practical model for super-microsurgery training using the middle caudal arteries of Kunming mice. Methods: A ⊔-shaped incision was made approximately 1 cm from the root of the tail in the mouse, and the skin, together with the subcutaneous tissue, was turned up into a rectangular shape to the opposite side with exposure of the mouse middle caudal artery and the accompanying veins. The artery was freed for approximately 1 cm in length. The middle caudal artery was cut transversely at the site, and then the severed middle caudal artery was anastomosed end-to-end using 12-0 microsutures in the order of 6, 12, 3, and 9 o'clock with four stitches. Results: The mouse caudal artery had a constant anatomical location accompanied by a vein. The immediate postoperative patency after vascular anastomosis was 100% (15/15) in all mouse models, the postoperative patency was 100% (5/5), 80% (4/5), and 75% (3/4) at 24 h, 3 days, and 1 week postoperatively, respectively. The outer diameter of the mouse middle caudal artery was 0.2 ~ 0.3 (0.22 ± 0.03) mm. The vascular anastomosis time was 6.5 ~ 15 (11.0 ± 2.5) min. Conclusion: The mouse middle caudal artery was superficially located and anatomically constant, making it easy to be located and exposed. The small size of the opening made it suitable for establishing a useful model for training in super-microsurgery vascular anastomoses.

Multifunctional biosensor constructed by Ag-coating magnetic-assisted unique urchin core porous shell structure for dual SERS enhancement, enrichment, and quantitative detection of multi-components inflammatory markers.

The simultaneous, precise, and quantitative detection of multi-components inflammatory markers (IMs) in sepsis serum by surface-enhanced Raman scattering (SERS) remains a challenging problem. A novel, multifunctional biosensor with dual enrichment and enhancement was designed for the ultrasensitive and quantitative analysis of multi-components IMs. The biosensor contains SERS tags-unique urchin core/porous shell (CPS) structure modified with Raman reporters (RaRs), magnetic assist-Ag coated Fe3O4 magnetic nanoparticles (Ag MNPs) modified with internal standard (IS), and then aptamer (Apt) modification to form the sandwich structure (Ag MNPs/IMs/CPS). This multifunctional sensor used for IMS detection has the following innovations: The intensity ratio IRaRs/IIS with Lg CIMs present a good and wide linear relationship to achieve the simultaneous, precise, and quantitative detection of IMS in serum; The detection results display ultrasensitivity, and the limit of detection (LOD) for CRP, IL-6, and PCT is 100 fg/mL, 0.1 fg/mL, and 1.0 fg/mL, which is lower than other detection techniques; The calculated data of clinical blood samples of sepsis by this SERS method is consistent with the hospital results, and can provide more compositional data of IMs. Thus, this combined approach developed a sensing platform for rapid screening, accurate evaluation, early warning, and diagnosis of sepsis.

Secondary-structure switch regulates the substrate binding of a YopJ family acetyltransferase.

The Yersinia outer protein J (YopJ) family effectors are widely deployed through the type III secretion system by both plant and animal pathogens. As non-canonical acetyltransferases, the enzymatic activities of YopJ family effectors are allosterically activated by the eukaryote-specific ligand inositol hexaphosphate (InsP6). However, the underpinning molecular mechanism remains undefined. Here we present the crystal structure of apo-PopP2, a YopJ family member secreted by the plant pathogen Ralstonia solanacearum. Structural comparison of apo-PopP2 with the InsP6-bound PopP2 reveals a substantial conformational readjustment centered in the substrate-binding site. Combining biochemical and computational analyses, we further identify a mechanism by which the association of InsP6 with PopP2 induces an α-helix-to-ß-strand transition in the catalytic core, resulting in stabilization of the substrate recognition helix in the target protein binding site. Together, our study uncovers the molecular basis governing InsP6-mediated allosteric regulation of YopJ family acetyltransferases and further expands the paradigm of fold-switching proteins.

Sizes and ligands tuned gold nanocluster acting as a new type of monoamine oxidase B inhibitor.

Monoamine oxidase inhibitors (MAOIs) are a class of drugs that can be used in the treatment of Parkinson's disease, clinical depression, and anxiety by targeting monoamine oxidase B (MAO). However, the side effects of MAOIs drive the requirement of a new framework of enzyme inhibitors development. In this context, a new type of MAOI has been built on the framework of gold nanoclusters (AuNCs), realizing the transformation from no function of small molecules to MAOI function of ligand-modified AuNCs. The MAOI activity of fabricated AuNCs can be achieved by size control and specific ligands modification. In this work, AuNCs modified with cysteamine or 4-aminothiophenol, about 1-3 nm in size, were found to have MAOI activity (MAOI-like AuNCs) and their characterization has been extensively described. Meanwhile, the possible mechanism behind this MAOI activity has been explored and it is believed that the proper size of AuNCs with ligands containing amino groups can bind tightly with the entrance to active sites of MAO, blocking the enzyme interacting with its substrates, thereby realizing the function of MAOI. Last, the antimicrobial activity and the performance of the MAOI-like AuNCs in the human blood sample were explored and suggested that MAOI-like AuNCs do not possess strong antimicrobial activity and have no visualized side effect on blood cells, although the by-product peroxide of MAO reaction may reshape the white blood cells. The research in this work may shed some light on the development of a new type of enzyme inhibitor based on the framework of nanomaterials.

Development of a tibial experimental non-union model in rats.

BACKGROUND: Many non-union animal models have been developed to explore the problems surrounding fracture healing. However, the existing models are not perfect and cannot satisfy all non-union studies. This study aimed to make a non-union model of the tibia in rats by cauterization of the posterior of 2 mm on both sides of the fracture end after open osteotomy of the tibia and fixing the fractured tibia with a Kirschner wire 0.8 mm in diameter. METHODS: For this study, 96 female adult Sprague-Dawley (SD) rats were used. The rats underwent surgery to produce a tibial open fracture and were fixed with a 0.8-mm diameter Kirschner wire. In 48 of the rats, the periosteum proximal and distal to the fracture end was cauterized. RESULTS: At 2, 4, 6, and 8 weeks after surgery, radiological and histological analysis showed typical physiological healing in the control group, and the healing rate was 100% at 6 weeks. But the non-union group was characterized by resorption of the fracture ends with few callus formations and no bridging callus formation, and the healing rate was 0% at 8 weeks. CONCLUSIONS: This method represents a reproducible model to create atrophic non-unions. This model provides a new option for studying the basic healing mechanisms and evaluating new therapies for bone regeneration and treatment of non-unions.

Reconstruction of Soft Tissue Defects in the Hand with a Free Anterolateral Thigh Deep Fascia Flap.

OBJECTIVE: To report our experience in the reconstruction of soft tissue defects in the hand with a free anterolateral thigh deep fascia flap and describe the clinical outcomes. METHODS: This study was a retrospective trial. From November 2016 to January 2020, six patients (four men and two women) with soft tissue defects in the hand were included in this study. The average age of the patients was 33.7 ± 12.7 years (range, 20 to 50 years). All patients underwent reconstructions with free anterolateral thigh deep fascia flaps. Relevant clinical characteristics were recorded prior to surgery. The size and thickness of the deep fascia flap and the thickness of the skin were measured intraoperatively. The survival of the flaps and skin grafts and the occurrence of infection were recorded after the operation. At follow-up, donor site complications and postoperative effects were evaluated according to the outcome satisfaction scale. The pain in the injured hand was assessed using the visual analog scale. RESULTS: The average body mass index (BMI) was 26.6 ± 1.7 kg/m2 (range, 23.9 to 28.7 kg/m2 ). The defect sizes ranged from 5 cm × 5 cm to 13 cm × 8 cm (average, 53.1 ± 27.9 cm2 ). The six anterolateral thigh deep fascia flaps ranged from 7 cm × 6 cm to 14 cm × 9 cm in size (average, 71.8 ± 29.1 cm2 ). The thicknesses of skin ranged from 25 mm to 40 mm (average, 32.5 ± 4.8 mm), and the thicknesses of the deep fascia flaps ranged from 2 mm to 3 mm (average, 2.5 ± 0.5 mm). After the operation, the blood supply of the deep fascia flap was normal in all cases. The second-stage skin grafts of most patients survived completely. The skin graft in one case was partially necrotic and healed after a dressing change. No infection occurred. At follow-up (average, 16.3 ± 6.9 months), there was only a linear scar and no loss of sensation at the donor site of each patient. According to the outcome satisfaction scale, the outcome satisfaction score ranged from 6 to 8 (average, 7.2 ± 0.9), all of which were satisfactory. Apart from one patient who reported mild pain, all the other patients reported no pain. Three typical cases are presented in this article. CONCLUSIONS: The free anterolateral thigh deep fascia flap, which is suitable for reconstruction of soft tissue defects in the hand, can provide very good outcomes both functionally and aesthetically.

Real-time monitoring of aristolochic acid I reduction process using surface-enhanced Raman Spectroscopy with DFT simulation.

With the increasing number of reports on aristolochic acid I (AAI), more and more toxic and side effects have been discovered successively. The main recognized carcinogenic mechanism is that AAI is metabolized into aristololactam I (AAT) in the body by nitroreductases, ultimately forming AAT-DNA adducts that cause disease. However, the carcinogenic mechanism is still not well understood by currently reported indirect method, there has always been a great demand to develop a direct method for real-time monitoring such process. In this work, surface-enhanced Raman spectroscopy (SERS) was used for the first time to monitor the process of AAI under the action of reducing agent sodium borohydride and catalyst Raney nickel to form AAT. We first found the abundant intermediate product-amino derivative of AAI, which was never reported before by other methods. The AAT was then obtained by a one-step dehydration reaction from the amino derivative of AAI under such reduction conditions. The product of amino derivative of AAI and AAT were further verified by thin-layer chromatography, H nuclear magnetic resonance spectra, mass spectrometry, and ultra-high performance liquid chromatography. Furthermore, a density functional theory-supported in-depth vibrational characterization of AAI and AAT was performed. The monitoring of the AAI reduction process by SERS can be of great significance for further exploration of its pathogenic mechanism, prevention, and monitoring of "nephropathy" and other diseases caused by AAI.

Click-Reaction-Triggered SERS Signals for Specific Detection of Monoamine Oxidase B Activity.

Human monoamine oxidases (MAOs) play important roles in maintaining the homeostasis of biogenic amines. One of its isoforms, monoamine oxidase B (MAOB), is thought to be involved in several neurodegenerative diseases, which make the selective detection of MAOB activity essential. In this work, a novel surface-enhanced Raman scattering (SERS) sensor was fabricated and the MAOB activity was specifically determined by detecting the SERS signals of an enzyme-catalyzed reaction product via an amine-aldehyde click reaction. This process was simply achieved by coating core-shell gold-silver nanoparticles (Au@Ag NPs) on 3-aminopropyl aminopropyl triethoxysilane (APTES)-modified glass, and then, a monolayer of cysteamine (CA) was attached to the nanoparticle surface as a linker through Ag-S bonds. Using phenethylamine (PA) as a specific substrate of MAOB, the enzyme product phenylacetaldehyde (PAA) will produce significant Raman signals via the amine-aldehyde click reaction with CA, while other molecules, such as MAOB and PA, have no signal output because they cannot form close interaction with nanoparticles due to the existence of a CA layer. This sensor was further used for the specific determination of MAOB activity in clinical blood samples and the MAOB inhibitor assessment successfully. Meanwhile, by changing the click reaction types and taking advantage of the SERS fingerprint peaks for the specific click reaction products, this strategy offers huge potential to detect multiple enzyme activities simultaneously and can be used for new click reaction screening, enzyme-related disease diagnosis, drug screening, and clinical diagnosis.

Ischemia Injury: A New Method Accelerates Bone Healing in a Rat Tibia Fracture Model.

To find a simple and noninvasive method to promote fracture healing, we are trying to explore whether repetitive brief ischemia would promote bone healing. 88 rats divided into 6 groups were used to make right tibia closed fracture caused by the heavy weight collision method. Healthy side groups received homemade tourniquet placed on left and affected side group placed on right thigh 10 min inflated/10 min deflated 3 times every 24 hours or 48 hours after tibia fractured. Rats in control groups received homemade tourniquet uninflated placed on right thigh 1 hour every 24 hours or 48 hours. X-rays were checked at 1, 2, and 4 weeks. Micro-CT inspected the bone healing at 2 and 4 weeks. Serum cytokines, such as bone morphogenetic protein-2 (BMP-2), vascular endothelial growth factor (VEGF), diethanolamine enzyme activity unit of alkaline phosphatase (ALP) and transforming growth factor-ß1 (TGF-ß1), interleukin 10 (IL-10) and interleukin 6 (IL-6), were checked at 1, 2, and 4 weeks. Local histology was evaluated at 2 weeks. HE dye and BMP-2, VEGF, TGF-ß, and ALP immunohistochemical stains were made. Callus areas of posterior-anterior and lateral views were calculated and repetitive brief ischemia increased the callus areas ratio at 1 and 2 weeks. Besides, from micro-CT results, repetitive brief ischemia increased the bone volume (BV) at 2 and 4 weeks and also increased the total bone tissue volume (TV) at 2 weeks and BV/TV at 4 weeks. The serum cytokines, such as BMP-2, VEGF, diethanolamine enzyme activity unit of ALP and TGF-ß1, have increased by repetitive brief ischemia at 1, 2 weeks. It is opposite of affected side group that the level of serum IL-10 increased and IL-6 decreased in healthy side group at 1, 2 weeks. Repetitive brief ischemia increased the callus area at 2 weeks and boosted the synthesis of BMP-2, VEGF, TGF-ß, and ALP in the fracture region at 2 weeks from tissue stains. Repetitive brief ischemia promotes bone healing no matter on the affected side or the healthy side limb.

The p38 MAPK Inhibitor SB203580 Abrogates Tumor Necrosis Factor-Induced Proliferative Expansion of Mouse CD4+Foxp3+ Regulatory T Cells.

There is now compelling evidence that tumor necrosis factor (TNF) preferentially activates and expands CD4+Foxp3+ regulatory T cells (Tregs) through TNF receptor type II (TNFR2). However, it remains unclear which signaling transduction pathway(s) of TNFR2 is required for the stimulation of Tregs. Previously, it was shown that the interaction of TNF-TNFR2 resulted in the activation of a number of signaling pathways, including p38 MAPK, NF-κB, in T cells. We thus examined the role of p38 MAPK and NF-κB in TNF-mediated activation of Tregs, by using specific small molecule inhibitors. The results show that treatment with specific p38 MAPK inhibitor SB203580, rather than NF-κB inhibitors (Sulfasalazine and Bay 11-7082), abrogated TNF-induced expansion of Tregs in vitro. Furthermore, upregulation of TNFR2 and Foxp3 expression in Tregs by TNF was also markedly inhibited by SB203580. The proliferative expansion and the upregulation of TNFR2 expression on Tregs in LPS-treated mice were mediated by TNF-TNFR2 interaction, as shown by our previous study. The expansion of Tregs in LPS-treated mice were also markedly inhibited by in vivo treatment with SB203580. Taken together, our data clearly indicate that the activation of p38 MAPK is attributable to TNF/TNFR2-mediated activation and proliferative expansion of Tregs. Our results also suggest that targeting of p38 MAPK by pharmacological agent may represent a novel strategy to up- or downregulation of Treg activity for therapeutic purposes.

A specific immune tolerance toward offspring cells is to exist after the mother lymphocyte infusion.

PURPOSE: To examine immune tolerance between maternal lymphocytes and offspring tissue after a donor lymphocyte infusion. METHODS: Mouse models were established by mating female BALB/c mice with male C57BL mice. Splenic lymphocytes from donors of different genetic backgrounds were labeled with carboxyfluorescein succinimidyl ester (CFSE), and 1×107 of the labeled cells were intravenously injected into a recipient. At 6h, 24h, 72h and 120h after the infusion, mononuclear cells in recipient spleen, liver, thymus, lymph nodes, and peripheral blood were collected. CFSE+, CFSE-, CD3+, CD8+, CD4+, CD19+, NK1.1+, CD25+, and CD127+ lymphocytes in those samples were analyzed by flow cytometry. The distribution of donor T cells, B cells, NK cells, helper T cells, cytotoxic T cells, and recipient regulatory T cells in the tissues were then analyzed. RESULTS: Maternal lymphocytes were more likely to survive in offspring. At 120h after infusion, the percentages of maternal cells in the offspring were 0.52±0.11% in lymph nodes, 0.97±0.04% in peripheral blood, and 0.97±0.11% in the spleen. Few donor cells, if any, were detected in these tissues at 120h after aunt to child, father to child, and unrelated allogeneic infusions were performed. The subtype proportion of donor lymphocytes changed significantly in the recipient tissues. Recipient Treg cells increased in the mother to child group, but not in the aunt to child, father to child, and unrelated allogeneic groups, suggesting a decreased cellular immune response to allogeneic cells in the mother to child group. At 120h after the infusion, no donor cells were detected in the recipient livers and thymuses of all groups, implying that donor cells were barely able to colonize in the liver and thymus. CONCLUSION: Specific immune tolerance to maternal lymphocytes exists in offspring. An infusion of maternal donor lymphocytes may produce a relatively persistent effect of adoptive immunotherapy with reduced side-effects.

[LNK Gene Single Nucleotide Polymorphisms and Acute Leukemia Susceptibility].

To investigate the relationship between the LNK(SH2B3) gene single nucleotide polymorphism and risk of acute leukemia (AL) in Chinese population. METHODS: The bone marrow and peripheral blood samples from 31 cases of acute lymphoblastic leukemia, 70 cases of acute myeloid leukemia and 130 healthy persons as the controls were collected. Genotype of LNK SNP Rs3184504(c.784T>C) and Rs78894077(c.724C>T) were determined by PCR-RFLP, and were confirmed by gel electrophoresis and sequencing. The NB4, THP-1 and Raji leukemia cell line models were cultured, the leukemia cell line LNK Rs3184504 and Rs78894077 polymorphism were detected by using direct sequencing. RESULTS: The CC genotype frequencies of Rs3184504 SNP were higher in ALL and AML patients than those in control (P<0.01), but there was no different between the groups in AML and ALL. The frequency of LNK gene Rs3184504 C allele was higher in AL as compared with control (P<0.01). The LNK gene Rs78894077 locus genotype distribution was not significantly different between the AL and the normal control group (P>0.05). Both Rs3184504 and Rs78894077 sites were detected as CC genotype in NB4, THP-1 and Raji cells. CONCLUSION: The persons carrying C allele of LNK gene Rs3184504 are more prone to develop acute leukemia.