P38γ modulates the lipid metabolism in non-alcoholic fatty liver disease by regulating the JAK-STAT signaling pathway.

Non-alcoholic fatty liver disease (NAFLD) is a major health problem in Western countries and has become the most common cause of chronic liver disease. Although NAFLD is closely associated with obesity, inflammation, and insulin resistance, its pathogenesis remains unclear. The disease begins with excessive accumulation of triglycerides in the liver, which in turn leads to liver cell damage, steatosis, inflammation, and so on. P38γ is one of the four isoforms of P38 mitogen-activated protein kinases (P38 MAPKs) that contributes to inflammation in different diseases. In this research, we investigated the role of P38γ in NAFLD. In vivo, a NAFLD model was established by feeding C57BL/6J mice with a methionine- and choline-deficient (MCD) diet and adeno-associated virus (AAV9-shRNA-P38γ) was injected into C57BL/6J mice by tail vein for knockdown P38γ. The results indicated that the expression level of P38γ was upregulated in MCD-fed mice. Furthermore, the downregulation of P38γ significantly attenuated liver injury and lipid accumulation in mice. In vitro, mouse hepatocytes AML-12 were treated with free fatty acid (FFA). We found that P38γ was obviously increased in FFA-treated AML-12 cells, whereas knockdown of P38γ significantly suppressed lipid accumulation in FFA-treated AML-12 cells. Furthermore, P38γ regulated the Janus Kinase-Signal transducers and activators of transcription (JAK-STAT) signaling pathway. Inhibition of P38γ can inhibit the JAK-STAT signaling pathway, thereby inhibiting lipid accumulation in FFA-treated AML-12 cells. In conclusion, our results suggest that targeting P38γ contributes to the suppression of lipid accumulation in fatty liver disease.

Cuproptosis: unveiling a new frontier in cancer biology and therapeutics.

Copper plays vital roles in numerous cellular processes and its imbalance can lead to oxidative stress and dysfunction. Recent research has unveiled a unique form of copper-induced cell death, termed cuproptosis, which differs from known cell death mechanisms. This process involves the interaction of copper with lipoylated tricarboxylic acid cycle enzymes, causing protein aggregation and cell death. Recently, a growing number of studies have explored the link between cuproptosis and cancer development. This review comprehensively examines the systemic and cellular metabolism of copper, including tumor-related signaling pathways influenced by copper. It delves into the discovery and mechanisms of cuproptosis and its connection to various cancers. Additionally, the review suggests potential cancer treatments using copper ionophores that induce cuproptosis, in combination with small molecule drugs, for precision therapy in specific cancer types.

Perception of Perioperative Risk for Arthroplasty Patients: A Poll of Chinese Orthopaedic Surgeons.

BACKGROUND: Informed by the precedent of an American Association of Hip and Knee Surgeons (AAHKS) survey, where 95% of participants reported instigating modifications to preoperative risk factors, this study appraised the approach of Chinese arthroplasty surgeons toward patients who had modifiable risks. METHODS: An adaptation of the AAHKS survey tool for a Chinese cohort was undertaken. The survey queried 600 Chinese Society of Hip and Knee Surgeons members on whether the perception of unoptimized medical comorbidities and socioeconomic elements affects the propensity to offer surgical procedures. RESULTS: Out of the distributed surveys, 150 responses were received, culminating in a response rate of 25%. The data illustrate that 98.7% of Chinese surgeons practice restrictions on arthroplasty access for patients who have modifiable risk factors, with notable frequencies for malnutrition (93.3%), anemia (91.3%), recent hyaluronic acid injections (within one month, 88.7%), and corticosteroid injections (within 3 months, 74.7%). Assessment criteria ahead of surgery included limitations such as a body mass index under 40 (47.3%), requirements for smoking cessation (57.3%), an acceptable hemoglobin A1c level (95.3%), and a dependent fasting blood glucose level (88%). Moreover, 87.3% of respondents endorsed the need for additional interventions for certain socioeconomically disadvantaged patients to achieve successful outcomes. A majority of respondents (94.7%) believed that more equitable access to care, facilitated by better-adjusted payment methodologies, could enhance patient outcomes. Current payment schemes were also perceived by a majority to potentially compromise outcomes for patients who have inadequate social support (80.7%), low socioeconomic status (67.3%), and those lacking insurance (72.7%). CONCLUSIONS: The consistency of the almost 99% response rate in addressing modifiable risk factors prior to arthroplasty aligns closely with the reported practices in the AAHKS survey. These findings underscore the shared valuation of preoperative risk factor optimization by Chinese and American arthroplasty surgeons, notwithstanding divergent healthcare system structures.

Eupatilin alleviates inflammatory response after subarachnoid hemorrhage by inhibition of TLR4/MyD88/NF-κB axis.

Early brain injury (EBI) is associated with the adverse prognosis of subarachnoid hemorrhage (SAH) patients. The key bioactive component of the Chinese herbal medicine Artemisia asiatica Nakai (Asteraceae) is eupatilin. Recent research reports that eupatilin suppresses inflammatory responses induced by intracranial hemorrhage. This work is performed to validate whether eupatilin can attenuate EBI and deciphers its mechanism. A SAH rat model was established by intravascular perforation in vivo. At 6 h after SAH in rats, 10 mg/kg eupatilin was injected into the rats via the caudal vein. A Sham group was set as the control. In vitro, BV2 microglia was treated with 10 µM Oxyhemoglobin (OxyHb) for 24 h, followed by 50 µM eupatilin treatment for 24 h. The SAH grade, brain water content, neurological score, and blood-brain barrier (BBB) permeability of the rats were measured 24 h later. The content of proinflammatory factors was detected via enzyme-linked immunosorbent assay. Western blot analysis was conducted to analyze the expression levels of TLR4/MyD88/NF-κB pathway-associated proteins. In vivo, eupatilin administration alleviated neurological injury, and decreased brain edema and BBB injury after SAH in rats. Eupatilin markedly reduced the levels of interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor-α (TNF-α), and suppressed the expression levels of MyD88, TLR4, and p-NF-κB p65 in the SAH rats' cerebral tissues. Eupatilin treatment also reduced the levels of IL-1ß, IL-6, and TNF-α, and repressed the expression levels of MyD88, TLR4, and p-NF-κB p65 in OxyHb-induced BV2 microglia. Additionally, pyrrolidine dithiocarbamate or resatorvid enhanced the suppressive effects of eupatilin on OxyHb-induced inflammatory responses in BV2 microglia. Eupatilin ameliorates SAH-induced EBI via modulating the TLR4/MyD88/NF-κB pathway in rat model.

Plantar Pressure Distribution and Posture Balance During Walking in Individuals with Unilateral Chronic Ankle Instability: An Observational Study.

BACKGROUND Patients with chronic ankle instability (CAI) can present with abnormal gait. The purpose of this study was to evaluate plantar pressure distributions and posture balance during walking in unilateral CAI patients. MATERIAL AND METHODS We recruited 24 unilateral CAI patients and 24 healthy individuals; plantar pressure analysis was conducted using the Footscan® 3D pressure system. The following parameters were assessed and recorded: peak force/weight (PF/W), time to peak force (TPF), time to boundary (TTB), and COP velocity. The differences between the affected and unaffected side of the CAI group and control group were determined. Pearson correlation analysis and univariate analysis was used to investigate the correlation between plantar pressure parameters and related factors. RESULTS The comparison of PF/W showed that the plantar pressure of both sides in the CAI group were laterally distributed. The comparison of TPF, TTB, and COP velocity in different groups showed that the posture balance on the affected side of CAI patient was more impaired than the unaffected side and the control group. Male patients with CAI tend to have better posture balance than females, and a low CAIT score is correlated with poor posture balance. CONCLUSIONS The plantar pressure on both sides in unilateral CAI patients was laterally distributed and their balance function was impaired. It is necessary for CAI patients to receive functional training of both sides during rehabilitation, and plantar pressure analysis is promising for diagnosis and evaluation of CAI.

Microsurgical Management of Fourth Ventricle Lesions Via the Median Suboccipital Keyhole Telovelar Approach.

In this 2-year retrospective analysis, 13 patients with fourth ventricle lesions who underwent microsurgical resection via the midline suboccipital keyhole telovelar approach were analyzed. This is the first study to investigate the surgical outcome and complications of using this approach to resect various types of lesions in the fourth ventricle. We aimed to clarify whether this approach has met its promise of lesion dissection. Three patients (23.1%) had intraoperative extraventricular drains. There were no immediate postoperative deaths. Gross total resection was achieved in 84.6% of the cases. The Fisher exact test showed there was no statistically significant correlation between lesion location, lesion size, brainstem invasion, and extent of resection. About two third (69.2%) of the cases were free of complications. New or worsening gait/focal motor disturbance (15.4%) was the most common neurological deficit in the immediate postoperative period. One patient (7.7%) had worse gait disturbance/motor deficit following surgical intervention. Two patients (15.4%) developed meningitis. Two patients (15.4%) required postoperative cerebrospinal fluid diversion after tumor resection, of these 2 patients, 1 (7.7%) eventually needed a permanent shunt. There were no cases of cerebellar mutism and bulbar paralysis. The median suboccipital keyhole telovelar approach provides relative wide access to resect most fourth ventricle tumors completely and with satisfactory results. In contrast, this requires the appropriate patient selection and skilled surgeons.

Site-Specific Synthesis of Protein-Oligo Conjugates through Histidine-Maleimide-Mediated Imidazolidinone Formation.

Protein modifications are crucial for biological applications including detection and therapeutics. Methods that site-specifically modify target proteins are limited and demanding. Here, we present a novel N-terminal labeling technique that improves not only the conjugation efficiency but also the ratio for mono-modification. This technique has been applied to modify multiple protein targets efficiently. In addition, this new linkage was found to be hydrolytically stable under extreme thermal pressure (95 °C heating under aqueous buffer). Of significant note, protein functions including binding and folding properties were retained after conjugation.

Tetrandrine Ameliorates Traumatic Brain Injury by Regulating Autophagy to Reduce Ferroptosis.

Traumatic brain injury (TBI) is the leading cause of death and disability in trauma patients. However, the effects and mechanism of autophagy after TBI remain unclear. This study aimed to investigate whether tetrandrine could ameliorate TBI through autophagy to reduce ferroptosis. A mice model for TBI was implemented. Behavioral and histomorphological experiments were performed to evaluate outcomes of the mice. The ferroptosis levels was detected by Perls staining. Enzyme-linked immunosorbent assay (ELISA) was applied to detect malondialdehyde (MDA), glutathione (GSH), and glutathione peroxidase 4 (GPX4) levels in the brain tissue. Western blot test was performed to detect Beclin 1, light chain 3 (LC3) II/I, p62, GPX4, SCL7A11, and ferritin heavy chain 1 (FTH1) levels, and the expression of LC3B, Beclin 1, GPX4, and FTH1 in the brain tissue was detected by immunofluorescence (IF). The behavioral and histomorphological results demonstrated that tetrandrine improved the neurological function and cerebral edema on days 1, 3, and 7 after TBI. The ELISA results suggested that tetrandrine reduced the MDA concentration and increased GSH concentration on days 1, 3, and 7 after TBI. IF staining and Perls staining reflected that tetrandrine promoted autophagy and inhibited ferroptosis on days 1, 3, and 7 after TBI, respectively. Tetrandrine further improved the neurological function, cerebral edema, autophagy, and ferroptosis on days 1, 3, and 7 after TBI after adding the autophagy inducer rapamycin. The effect of TET in alleviating TBI increased with the increase of time and dose. Tetrandrine ameliorated TBI by regulating autophagy to reduce ferroptosis, providing a new therapeutic strategy for TBI.

Transcription factor ZFHX3 regulates calcium influx in mammary epithelial cells in part via the TRPV6 calcium channel.

Zinc finger homeobox 3 (ZFHX3) is a transcription factor that regulates multiple cellular processes including cell proliferation, differentiation and neoplastic development. It is also involved in the function of steroid hormones estrogen and progesterone and the peptide hormone prolactin in mammary epithelial cells. In this study, we investigated whether and how ZFHX3 regulates intracellular calcium homeostasis in mammary epithelial cells. We found that ZFHX3 affected both store operated calcium entry and store independent calcium entry (SOCE and SICE). Simultaneously, the expression of the calcium channel TRPV6 was regulated by ZFHX3, as demonstrated by expression analysis and luciferase reporter assay. In cells with knockdown of ZFHX3, calcium entry was partially rescued by the overexpression of wild type but not the pore mutants of TRPV6. In addition, overexpression of TRPV6 promoted differentiation of the MCF10A mammary epithelial cells in three-dimensional culture, which is consistent with our previous findings that ZFHX3 is essential for mammary gland differentiation. These findings suggest that ZFHX3 plays an important role in intracellular calcium homeostasis in mammary epithelial cells, at least in part, by regulating TRPV6.

Resolving quantum dots and peptide assembly and disassembly using bending capillary electrophoresis.

Despite the numerous techniques developed for the studying nanoparticle and peptide interaction nowadays, sensitive and convenient assay in the process of flow, especially to simulate the self-assembly of quantum dots (QDs) and peptide inflow in blood vessels, still remains big challenges. Here, we report a novel assay for studying the self-assembly of QDs and peptide, based on CE using a bending capillary. We demonstrate that the semicircles numbers of the bending capillary affect the self-assembly kinetics of CdSe/ZnS QDs and ATTO-D3 LVPRGSGP9 G2 H6 peptide. Moreover, benefitting from this novel assay, the effect of the position on the self-assembly has also been realized. More importantly, we also demonstrate that this novel assay can be used for studying the stability of the QDs-peptide complex inflow. We believe that our novel assay proposed in this work could be further used as a general strategy for the studying nanoparticle-biomolecule interaction or biomolecule-biomolecule interaction.

Three-dimensional printing assisted transcatheter closure of atrial septal defect with deficient posterior-inferior rim.

OBJECTIVE: Though successful transcatheter closure has been reported in secundum atrial septal defect (ASD) with deficient posterior-inferior rim, it is still difficult to screen the appropriate candidates. Three-dimensional printing (3DP) makes in vitro trial occlusion possible, and might provide a feasible method in the prediction of successful closure. METHODS: Thirty-five consecutive ASD patients (10M/25F, age, 47.7 ± 11.8 years) with deficient posterior-inferior rim (≤3 mm) were referred for attempted transcatheter closure, and personalized heart model (elastic rubber) was produced based on end-systolic MSCT images. The in vitro measurement and trial occlusion were performed for preoperative evaluation (in vitro successful/unsuccessful group), and the results were compared with postoperative outcomes. RESULTS: Successful in vitro occlusion was achieved in 30 patients (7M/23F), and the size of ASD was 27.1 ± 4.4 mm. The posterior-inferior rim was 0.95 ± 1.22 mm (rim defect in 17 patients), and 12 patients were associated with aortic rim deficiency. The subsequent transcatheter closure was performed successfully in 29 patients, and the occluder-diameter was identical to that of in vitro occlusion (35.0 ± 4.4 mm). The follow-up (1.4 ± 0.58 years) showed no residual shunt and related-complications. In unsuccessful in vitro group (n = 5), the range of rim deficiency was wider (P = 0.019) and the rim to inferior vena cava was shorter (4.60 ± 2.07 mm vs. 10.71 ± 5.28 mm, P = 0.016). Furthermore, transcatheter closure failed in all patients. CONCLUSIONS: In ASD with deficient posterior-inferior rim, 3DP allows accurate determination of the size and surrounding rims of ASD. Based on personalized heart model, in vitro trial occlusion is an effective method to identify the appropriate candidates for transcatheter closure.

A CE-FL based method for real-time detection of in-capillary self-assembly of the nanoconjugates of polycysteine ligand and quantum dots.

Small molecules with free thiol groups always show high binding affinity to quantum dots (QDs). However, it is still highly challenging to detect the binding capacity between thiol-containing molecules and QDs inside a capillary. To conquer this limitation, a capillary electrophoresis with fluorescence detection (CE-FL) based assay was proposed and established to investigate the binding capacity between QDs and a poly-thiolated peptide (ATTO 590-DDSSGGCCPGCC, ATTO-C4). Interestingly, the results showed that interval time had a great influence on QDs and ATTO-C4 self-assembly, which can be attributed to longer interval time benefitting the binding of QDs to ATTO-C4. The stability assays on ATTO-C4-QD assembly indicated that high concentration of imidazole or GSH had a high capability of competing with the bound ATTO-C4, evidenced by dramatically dropping of S 625/S 565 ratio from 0.78 to 0.30 or 0.29. Therefore, all these results above suggested that this novel CE-FL based detection assay could be successfully applied to the binding studies between QDs and thiol-containing biomolecules.

Actinide embedded nearly planar gold superatoms: structural properties and applications in surface-enhanced Raman scattering (SERS).

Planarity is a special property of superatoms, different from atoms. In this work, we predicted a series of nearly planar structures, An@Au6 (An = Ac-1, Th, Pa+1) clusters, using density functional theory (DFT). Calculations of these actinide embedded clusters reveal a 10-electron (1s21p41d4) closed-shell singlet configuration. It is found that all An@Au6 clusters are nearly or purely planar structures with only in-plane two-dimensional occupied superatomic molecular orbitals (SAMOs). In addition, applying them as surface-enhanced Raman scattering (SERS) substrates, the charge-transfer (CT) states at 677 nm (1dmetal-π1*pyridine) can lead to a SERS signal enhancement of 104 for a pyridine-Th@Au6 complex. Our research indicates that actinide embedded nearly planar superatomic clusters have unique optical properties and potential application value.

Percutaneous CT-guided radiofrequency ablation for patients with extrahepatic oligometastases of hepatocellular carcinoma: long-term results.

BACKGROUND: The aim of this study was to evaluate the therapeutic outcome of percutaneous computed tomography (CT)-guided radiofrequency ablation (RFA) for extrahepatic oligometastases of hepatocellular carcinoma (HCC). METHODS: Institutional review board approval was obtained for this retrospective study, and all patients provided written informed consent. Between April 2004 and December 2015, 116 oligometastases (diameter, 5-50 mm; 20.3 ± 10.4) in 79 consecutive HCC patients (73 men and 6 women; average age, 50.3 years ±13.0) were treated with RFA. We focussed on patients with 1-3 extrahepatic metastases (EHM) confined to 1-2 organs (including the lung, adrenal gland, bone, lymph node and pleura/peritoneum) who were treated naïve with curative intent. Survival, technical success and safety were evaluated. The log-rank test and Cox proportional hazards regression models were used to analyse the survival data. RESULTS: No immediate technical failure occurred, and at 1 month, the technique effectiveness rate was determined to be 95.8%. After a median follow-up time of 28.0 months (range, 6-108 months), the 1-, 2- and 3-year overall survival (OS) rates were 91, 70 and 48%, respectively, with a median survival time of 33.5 months. Time to unoligometastatic progression (TTUP) of less than 6 months (p < 0.001) and a Child-Pugh score of more than 5 (p = 0.001) were significant indicators of shorter OS. The 1-, 2- and 3-year disease free survival (DFS) rates were 34, 21 and 8%, respectively, with a median DFS time of 6.8 months. DFS was better for those with lung metastases (p = 0.006). Major complication occurred in nine (9.5%, 9/95) RFA sessions without treatment-related mortality. CONCLUSIONS: CT-guided RFA for oligometastatic HCC may provide favourable efficacy and technical success with a minimally invasive approach.

A novel in-capillary assay for dynamically monitoring fast binding between antibody and peptides using CE.

Nowadays, despite numerous techniques available for the detection of antibody-peptide binding, sensitivity and detection limit inflow still remains a major challenge. Herein, we report a strategy for the detection of binding inflow of monoclonal anti-FLAG M2 antibody and 5,6-carboxyfluorescein-labeled FLAG tag in capillary. Antibody and peptides were sequentially injected into the capillary where they mixed and bound together. Capillary electrophoresis with fluorescence detection was employed to monitor the binding process, and the results showed that the efficacy of the antibody-peptide binding in capillary (in-capillary assay) is affected by the stoichiometry. Compared to the out-capillary assay, this novel assay showed different KD values and required a very short detection time, thus showing great potential for rapid detection as well as other applications. Additionally, our novel assay can be used to investigate the stability of the antibody-peptide complex. The addition of excess DYKDDDDK or TAMRA-DYKDDDDK, with similar binding priorities with M2, can leads to dissociation of the complex with a two-step mechanism including dissociation and association. We believed that our developed method can be extended to monitor wide range of other biomolecule interactions.

Developing a capillary electrophoresis based method for dynamically monitoring enzyme cleavage activity using quantum dots-peptide assembly.

Herein, a novel assay has been developed for monitoring PreScission protease (His-PSP) mediated enzyme cleavage of ATTO 590 labeled peptide substrate (ATTO-LEV). This novel method is based on combining the use of capillary electrophoresis and fluorescence detection (CE-FL) to dynamically monitor the enzyme cleavage activity. A multivalent peptide substrate was first constructed by immobilizing His-tagged ATTO 590 labeled peptide substrate (ATTO-LEVH6) onto the surface of CdSe/ZnS quantum dots (QDs). Once successfully immobilized, the novel multivalent peptide substrate resulted in the Förster resonance energy transfer (FRET) from QDs to ATTO 590. The ATTO-LEVH6-QD assembly was then incubated with His-PSP to study the proteolytic cleavage of surface bound ATTO-LEVH6 by CE-FL. Our data suggests that PreScission-mediated proteolytic cleavage is enzyme concentration- and incubation time-dependent. By combining capillary electrophoresis, QDs and FRET, our study herein not only provides a new method for the detection and dynamically monitoring of PSP enzyme cleavage activity, but also can be extended to the detection of many other enzymes and proteases.

EGFL7 participates in regulating biological behavior of growth hormone-secreting pituitary adenomas via Notch2/DLL3 signaling pathway.

Growth hormone-secreting pituitary adenoma accounts for about 20% of the third most common intracranial neoplasm-pituitary adenomas-which makes up 15% of all intracranial tumors. The growth hormone-secreting pituitary adenoma invasion is a key risk factor associated with the operation results and highly correlated with the clinical prognosis. The epidermal growth factor-like domain multiple 7 protein, a unique 29 kDa secreted angiogenic factor, can result in pathologic angiogenesis and enhance the tumor migration and invasion. In this study, for the first time we found that epidermal growth factor-like domain multiple 7 protein expression was markedly higher in invasive growth hormone-secreting pituitary adenoma than non-invasive growth hormone-secreting pituitary adenoma. The tumor volume, histologic subtypes, invasiveness and recurrence of growth hormone-secreting pituitary adenoma were significantly associated with epidermal growth factor-like domain multiple 7 protein expression. Furthermore, we discovered that the histological classification methods of growth hormone-secreting pituitary adenoma according to electron microscopic examination and biological marker classification methods according to epidermal growth factor-like domain multiple 7 protein expression are more valuable in clinical application than the traditional classification methods based on Knosp and Hardy-Wilson grades. In summary, our results indicated epidermal growth factor-like domain multiple 7 protein participates in growth hormone-secreting pituitary adenoma proliferation and invasion regulation via Notch2/DLL3 signaling pathway. These findings raised the possibility that epidermal growth factor-like domain multiple 7 protein might serve as a useful biomarker to assess growth hormone-secreting pituitary adenoma invasion and prognosis or a potential therapeutic target for growth hormone-secreting pituitary adenoma treatment.

Multi-micronutrient supplementation during pregnancy for prevention of maternal anaemia and adverse birth outcomes in a high-altitude area: a prospective cohort study in rural Tibet of China.

Anaemia during pregnancy, characterised by Hb <110 g/l, is a specific risk factor for adverse maternal and perinatal outcomes in developing countries. The objective of this study was to determine the effectiveness of daily antenatal supplementation with multiple micronutrients (MMN) compared with folic acid (FA) on the occurrence of anaemia among pregnant women and their infants' health in a high-altitude area. A prospective cohort study was carried out in two rural counties in Tibet from 2007 to 2012. A total of 1149 eligible pregnant women were allocated daily supplementation with FA in one county and MMN containing a recommended allowance of twenty-three vitamins and minerals in another county starting ≤24 weeks of gestation and continuing until delivery. Compared with the FA group, prenatal supplementation with MMN was significantly associated with reduced odds of anaemia in the third trimester. This was demonstrated in the primary outcome, with an adjusted OR (AOR) of 0·63; 95 % CI 0·45, 0·88 and P=0·007 and also reduced odds of preterm delivery (AOR: 0·31; 95 % CI 0·15, 0·61; P=0·001). There was no difference between MMN and FA groups in mean birth weight (adjusted mean difference: 36·78; 95 % CI -19·42, 92·98 g; P=0·200), whereas MMN supplementation significantly reduced the odds of low-birth weight (LBW) babies (AOR: 0·58; 95 % CI 0·36, 0·91; P=0·019). In conclusion, the antenatal MMN supplementation in rural Tibet is associated with a reduction of maternal anaemia in the third trimester, and may potentially decrease the risk of preterm delivery and LBW babies.

Novel application of fluorescence coupled capillary electrophoresis to resolve the interaction between the G-quadruplex aptamer and thrombin.

The dynamic binding status between the thrombin and its G-quadruplex aptamers and the stability of its interaction partners were probed using our previously established fluorescence-coupled capillary electrophoresis method. A 29-nucleic acid thrombin binding aptamer was chosen as a model to study its binding affinity with the thrombin ligand. First, the effects of the cations on the formation of G-quadruplex from unstructured 29-nucleic acid thrombin binding aptamer were examined. Second, the rapid binding kinetics between the thrombin and 6-carboxyfluorescein labeled G-quadruplex aptamer was measured. Third, the stability of G-quadruplex aptamer-thrombin complex was also examined in the presence of the interfering species. Remarkably, it was found that the complementary strand of 29-nucleic acid thrombin binding aptamer could compete with G-quadruplex aptamer and thus disassociated the G-quadruplex structure into an unstructured aptamer. These data suggest that our in-house established fluorescence-coupled capillary electrophoresis assay could be applied to binding studies of the G-quadruplex aptamers, thrombin, and their ligands, while overcoming the complicated and costly approaches currently available.

In-capillary probing of quantum dots and fluorescent protein self-assembly and displacement using Förster resonance energy transfer.

Herein, a Förster resonance energy transfer system was designed, which consisted of CdSe/ZnS quantum dots donor and mCherry fluorescent protein acceptor. The quantum dots and the mCherry proteins were conjugated to permit Förster resonance energy transfer. Capillary electrophoresis with fluorescence detection was used for the analyses for the described system. The quantum dots and mCherry were sequentially injected into the capillary, while the real-time fluorescence signal of donor and acceptor was simultaneously monitored by two channels with fixed wavelength detectors. An effective separation of complexes from free donor and acceptor was achieved. Results showed quantum dots and hexahistidine tagged mCherry had high affinity and the assembly was affected by His6 -mCherry/quantum dot molar ratio. The kinetics of the self-assembly was calculated using the Hill equation. The microscopic dissociation constant values for out of- and in-capillary assays were 10.49 and 23.39 µM, respectively. The capillary electrophoresis with fluorescence detection that monitored ligands competition assay further delineated the different binding capacities of histidine containing peptide ligands for binding sites on quantum dots. This work demonstrated a novel approach for the improvement of Förster resonance energy transfer for higher efficiency, increased sensitivity, intuitionistic observation, and low sample requirements of the in-capillary probing system.