[Brand-New Ge20Ga5Sb10S65 Prism Biosensor Based on Inverted SPR].

Based on inverted surface plasmon resonance (ISPR) a novel biosensor consisting of Ge20Ga5Sb10S65-palladium-graphene layer-biomolecule layer is proposed. The refractive index of biomolecule layer alters as biomolecule experience interactions, thus leading to a shift of ISPR angle. On this basis, the spectrum output of sensor is derived by transfer matrix method. The sensitivity, the resolution, the dynamic detection range and the signal to noise ratio of the presented sensor are discussed and compared with the performance of traditional sensors. Moreover, the influences of grapheme layer thickness on sensors are analyzed with comparative study. Finally, near infrared is used as the incident light of the presented sensor. The results show that, the best thickness of grapheme layer is monolayer; the peak intensity of the ISPR reflection is about 80%~90% of intensity of incident light, guaranteeing a high signal to noise ratio; In the visible light, when λ = 632.8 nm, the presented sensor is 1.9 times the resolution of the sensor based on SiO2 coupling inverted surface plasmon resonance, is 3. 5 times the resolution of the sensor based on surface plasmon resonance(SPR), and is 2 times the dynamic detection range of pre-existing biosensor based on SPR. The application of Ge20Ga5Sb10S65 prism extends the detection light wavelength from the visible region to the near infrared region. When λ = 1,000 nm, the sensor is 3-4 times of the sensor in visible region. The research greatly contributes to the realization and application of biosensor based on inverted surface plasmon resonance.

Optimization of PROTAC Ternary Complex Using DNA Encoded Library Approach.

The proteolysis targeting chimera (PROTAC) strategy results in the down-regulation of unwanted protein(s) for disease treatment. In the PROTAC process, a heterobifunctional degrader forms a ternary complex with a target protein of interest (POI) and an E3 ligase, which results in ubiquitination and proteasomal degradation of the POI. While ternary complex formation is a key attribute of PROTAC degraders, modification of the PROTAC molecule to optimize ternary complex formation and protein degradation can be a labor-intensive and tedious process. In this study, we take advantage of DNA-encoded library (DEL) technology to efficiently synthesize a vast number of possible PROTAC molecules and describe a parallel screening approach that utilizes DNA barcodes as reporters of ternary complex formation and cooperative binding. We use a designed PROTAC DEL against BRD4 and CRBN to describe a dual protein affinity selection method and the direct discovery of novel, potent BRD4 PROTACs that importantly demonstrate clear SAR. Such an approach evaluates all the potential PROTACs simultaneously, avoids the interference of PROTAC solubility and permeability, and uses POI and E3 ligase proteins in an efficient manner.

[Up-regulation of the sHLA-G5 level by human-mouse chimeric anti-IL-2R monoclonal antibody treatment in kidney transplantation].

OBJECTIVE: To find whether the up-regulation of soluble human leucocyte antigen-G5 (sHLA-G5) levels is a new function mechanism of anti-interleukin-2 receptors (anti-IL-2R) monoclonal antibody treatment in kidney transplantation. METHODS: A total of 215 recipients at our centre from January 2006 to December 2007 were divided into antibody use group (n = 141) and antibody non-use group (n = 74) and another healthy group (n = 69). The sHLA-G5 level in peripheral blood was detected by enzyme-linked immunosorbent assay (ELISA). And the expression of HLA-G5 was confirmed by Western blot and Real-time polymerase chain reaction (PCR). RESULTS: sHLA-G5 levels was (56 ± 30) µg/L in using anti-IL-2 receptor monoclonal antibody before transplantation, It was higher than that before use antibody [(34 ± 20) µg/L], also higher than healthy group [(35 ± 17) µg/L] and antibody non-use group [(36 ± 19) µg/L, P < 0.05, respectively]. At Day 1, Day 4, Week 1, Week 2 post-transplantation, the level of sHLA-G5 of recipients with antibody use was significantly higher than that of those with antibody non-use. The values were as follows: (95 ± 35) µg/L vs (54 ± 16) µg/L, (131 ± 24) µg/L vs (75 ± 22) µg/L, (167 ± 44) µg/L vs (62 ± 17) µg/L, (172 ± 35) µg/L vs (45 ± 16) µg/L (all P < 0.01). And the results of Western blot and RT-PCR corresponded to those of ELISA. CONCLUSION: The preoperative use of first dose of anti-IL-2R monoclonal antibodies results in the up-regulated level of sHLA-G5. Thus it is beneficial for protecting the kidney survival and reducing the risks of acute rejection.

Serum fibroblast growth factor 23 for early detection of acute kidney injury in critical illness.

BACKGROUND: Serum fibroblast growth factor 23 (FGF23) is associated with acute kidney injury (AKI) and mortality in patients with critical illnesses. However, the accurate predictive performance of FGF23 on AKI remains inconclusive. METHODS: Meta-analysis was performed using data sources including PubMed, Web of Science, EMBASE, and Cochrane (until June 1, 2021). Cohort or observational studies including patients with AKI and serum FGF23 level as the index test were included. The primary outcome was the AKI detective accuracy. This study has been registered in PROSPERO (CRD42021249930). RESULTS: Eleven studies with 1946 patients in seven countries were included. Across all settings, the sensitivity and specificity for serum FGF23 levels to predict AKI were 82% (95% CI, 66-91%) and 77% (95% CI, 67-85%), respectively. The diagnostic odds ratio of FGF23 was 15.51 (95% CI, 4.89-49.19), with the pooled positive likelihood ratio of 3.62 (95% CI, 2.25-5.83) and a negative likelihood ratio of 0.23 (95% CI, 0.11-0.50). The area under the receiver operating characteristic curve to detect AKI was 0.86 (95% CI, 0.82-0.88). C-terminal FGF23 had a better performance than intact FGF23. CONCLUSIONS: Plasma FGF23 is a valuable biomarker for incident AKI in critically ill patients. Comparisons of FGF23 with other biomarkers in AKI still need more studies to prove.

[Applications of monitoring human leukocyte antigen G and its inhibitory receptors in post-renal transplantation].

OBJECTIVE: to study the feasibility of human leucocyte antigen-G (HLA-G) as a post-transplantation prognostic biomarker and discuss the correlation of its receptor expression and the mechanisms. METHODS: a total of 215 recipients in our centre from February 2006 to June 2008 were divided into stable kidney function group (n = 173) and acute rejection group (n = 42). The soluble human leucocyte antigen-G5 (sHLA-G5) level in peripheral plasma was detected by ELISA. And the HLA-G receptor ILT-2, KIR2DL4 on T, B, NK lymphocytes were analyzed by flow cytometry (FCM). The sHLA-G5 cutoff level by ROC curve was employed to predict the events of acute post-transplantation rejection. And regression analysis was used to determine the association of sHLA-G5 with acute rejection. RESULTS: an optimal cutoff value of 139.0 microg/L could be defined for sHLA-G5 (sensitivity: 63.6%, specificity: 82.1%, AUC: 0.780). Binary regression analysis showed that sHLA-G5 played an independent role on acute rejection (P = 0.019, OR = 0.039, 95%CI: 2.091 - 5.661). The rate of HLA-G receptor ILT-2 on CD4(+)T cell, CD8(+)T cell and B cell in acute rejection group was statistically lower than that in stable kidney function group (21% ± 7% vs 52% ± 17%, 23% ± 6% vs 39% ± 16%, 21% ± 7% vs 39% ± 16%, all P < 0.05). The expression of KIR2DL4 on NK cells in acute rejection group was statistically lower than that in stable kidney function group (31% ± 10%vs 57% ± 21%, P < 0.05). CONCLUSION: sHLA-G5 level may be predicted for acute rejection with a high sensitivity and specificity. The up-regulated expression of ILT-2 and KIR2DLT may contribute to immunology tolerance in peripheral circulation.

[Expression of human leucocyte antigen G in peripheral blood of kidney transplantation recipients].

OBJECTIVE: To investigate the expression of non-classical major histocompatibility complex (MHC)-I molecule, human leucocyte antigen (HLA) G, including membrane-bound HLA-G (mHLA-G), intracellular HLA-G (iHLA-G) and soluble HLA-G (sHLA-G), in peripheral blood of surviving kidney transplantation recipients and understand the relevance between HLA-G and the function of transplanted organ, as well as the onset of acute rejection. METHODS: A longitudinal study was performed on 175 kidney transplantation recipients. Three groups were involved in this study, including acute rejection group (n = 36), function stable group (n = 139) and healthy control group (n = 30). The expression of mHLA-G1 and iHLA-G1 in the T lymphocytes of peripheral blood was detected by flow cytometry analysis and the sHLA-G5 level detected by ELISA. RESULTS: The average rate of CD4(+)mHLA-G1(+), CD8(+)mHLA-G1(+), CD4(+)iHLA-G1(+), CD8(+)iHLA-G1(+) in T lymphocytes of healthy control group was 0.43% +/- 0.19%, 1.23% +/- 0.41%, 27% +/- 13% and 36% +/- 14% respectively. That of acute rejection group was 0.57% +/- 0.34%, 1.31% +/- 0.56%, 26% +/- 8% and 37% +/- 17%; that of function stable group was 0.61% +/- 0.43%, 1.39% +/- 0.47%, 26% +/- 9% and 37% +/- 17% respectively. There was no significant difference among the three groups (all P > 0.05). The average of sHLA-G5 levels in plasma of control group was (25 +/- 14) ng/ml, acute rejection group (24 +/- 15) ng/ml (pre-operative) and (34 +/- 21) ng/ml (post-operative), function stable group (25 +/- 11) ng/ml (pre-operative) and (56 +/- 32) ng/ml (post-operative). There was no significant difference among the three groups (pre-operative, P > 0.05). The average of sHLA-G5 levels in plasma of function stable group was higher than that of acute rejection group (post-operative, P < 0.05). CONCLUSION: There is a subset of CD4(+)HLA-G1(+) and CD8(+)HLA-G1(+)T lymphocytes with low percentage in peripheral blood of those surviving kidney transplantation recipients. The expressions of mHLA-G1 and iHLA-G1 have no relevance with the onset of acute rejection. sHLA-G5 is correlated with acute rejection in peripheral blood of surviving transplantation recipients.

Optimized methods for IL-17A refolding and anti-IL17A Fab production for co-crystallization with small molecules.

Refolding of human interleukin 17A (IL-17A) has been reported; however, the key refolding protocol was not robust enough to deliver consistent results and to be easily scaled up for crystallization. Here we report an optimized refolding method for IL-17A. Although co-crystal structures of IL-17A with ligands have been obtained with a high-affinity peptide and an anti-IL-17A Fab as stabilizers, neither the production yield nor the characterization of the IL-17A/Fab complex was reported. To facilitate co-crystallization of IL-17A with small-molecule compounds derived from our DNA encoded library, we also describe the method for yield enhancement of anti-IL-17A Fab production and characterize the IL-17A/Fab complex for the first time, providing an essential prerequisite for structure-based drug discovery targeting IL-17A.

Characterization of Specific N-α-Acetyltransferase 50 (Naa50) Inhibitors Identified Using a DNA Encoded Library.

Two novel compounds were identified as Naa50 binders/inhibitors using DNA-encoded technology screening. Biophysical and biochemical data as well as cocrystal structures were obtained for both compounds (3a and 4a) to understand their mechanism of action. These data were also used to rationalize the binding affinity differences observed between the two compounds and a MLGP peptide-containing substrate. Cellular target engagement experiments further confirm the Naa50 binding of 4a and demonstrate its selectivity toward related enzymes (Naa10 and Naa60). Additional analogs of inhibitor 4a were also evaluated to study the binding mode observed in the cocrystal structures.

Molecular mechanisms underlying the flavonoid-induced inhibition of alpha-synuclein fibrillation.

The molecular mechanism underlying the flavonoid-induced inhibition of alpha-synuclein fibrillation was thoroughly examined by various biochemical and biophysical approaches. The noncovalent binding of the inhibitory flavonoids to alpha-synuclein and the covalent modification by the flavonoid quinone led to the restriction of the conformational changes in this natively unfolded protein and to the stabilization of soluble flavonoid-modified species of alpha-synuclein (monomers and oligomers). All of these factors rather than a single one contribute to the inhibition of WT alpha-synuclein fibrillation induced by the flavonoid. The structural requirements that appear necessary to provide a flavonoid the ability to inhibit alpha-synuclein fibrillation were determined to be vicinal dihydroxyphenyl moieties, irrespective of the ring position where they are located. Flavonoids with three vicinal hydroxyl groups exhibited enhanced inhibitory effects on alpha-synuclein fibrillation. The antioxidant activities of flavonoids were generally correlated with their in vitro inhibitory effects on alpha-synuclein fibrillation. The flavonoids inhibiting alpha-synuclein fibrillation and stabilizing the protein monomeric conformation can serve as a model for the development of therapeutic drugs in combating Parkinson's disease.

HLA-G expression in the peripheral blood of live kidney transplant recipients.

BACKGROUND: The human leukocyte antigen-G (HLA-G) has been considered to be an important tolerogeneic molecule playing an essential role in maternal-fetal tolerance, upregulated in the context of transplantation, malignancy, and inflammation, and has been correlated with various clinical outcomes. The aim of this study was to investigate the clinical relevance of the expression of membrane HLA-G (mHLA-G), intracellular HLA-G (iHLA-G), and soluble HLA-G (sHLA-G) in the peripheral blood of live kidney transplant recipients. METHODS: We compared the expression of the three HLA-G isoforms in three groups, healthy donors (n=20), recipients with acute rejection (n=19), and functioning transplants (n=30). Flow cytometry was used to detect the expression of mHLA-G and iHLA-G in the T lymphocytes of peripheral blood from subjects in the three groups. Enzyme-linked immunosorbent assays were used to detect sHLA-G in the plasma from the three groups. RESULTS: There were no significant differences in mHLA-G and intracellular HLA-G among the three groups, but the sHLA-G plasma level was higher in the functioning group than in the acute rejection or healthy group. We found a subset of CD4(+)HLA-G(+) and CD8(+)HLA-G(+) T lymphocytes with low rates of mHLA-G expression in the peripheral blood of kidney transplantation recipients. Intracellular expression of HLA-G was detected in T lymphocytes. However, there was no correlation between acute rejection and the mHLA-G or intracellular HLA-G expression. CONCLUSION: sHLA-G was the major isoform in the peripheral blood of live kidney transplant recipients and high sHLA-G levels were associated with allograft acceptance.

[Impact of the land-use change on the non-point source nitrogen load in Yunmeng Lake watershed].

Take potable water sources in Linyi City Yunmeng Lake watershed as a case study, it obtains the nutrient export coefficient of land use by the export coefficient model and simulative rainfall experiment. On the basis of GIS and RS, it analyses the effect of the non-point source (NPS) pollution load because of the land-use change during the past 25 years. The result indicates that the TN increased from 3.77 x 10(3) t in 1986, to 4.45 x 10(3) t in 1995, to 5.5 x 10(3) t in 2010; As far as land-use type is concerned, the TN from farm-land increased year by year, the contribution rate is 80.11% in 1986, 82.60% in 1995 and 85.59% in 2010, the forestland and the grass-land load have a little change, but the contribution rate decreased gradually, the residential load increased by a large margin, however, the contribution rate is very little. As for the sub-basin, the higher the proportion of the farm-land is, the more the TN load increased. There is a significant positive correlation between the farm-land and the nitrogen (TN) load, so the farm-land is the sources of the nitrogen. Conversely, there are negative correlations between the forest-land, grass-land and the TN load; therefore, the forest-land and grass-land are the sinks of the nitrogen. Therefore, it can adjust the land-use structure to reduce and control the TN loss to water environmental pollution.

Effects of Various Flavonoids on the α-Synuclein Fibrillation Process.

α-Synuclein aggregation and fibrillation are closely associated with the formation of Lewy bodies in neurons and are implicated in the causative pathogenesis of Parkinson's disease and other synucleinopathies. Currently, there is no approved therapeutic agent directed toward preventing the protein aggregation, which has been recently shown to have a key role in the cytotoxic nature of amyloidogenic proteins. Flavonoids, known as plant pigments, belong to a broad family of polyphenolic compounds. Over 4,000 flavonoids have been identified from various plants and foodstuffs derived from plants and have been demonstrated as potential neuroprotective agents. In this study 48 flavonoids belonging to several classes with structures differing in the position of double bonds and ring substituents were tested for their ability to inhibit the fibrillation of α-synuclein in vitro. A variety of flavonoids inhibited α-synuclein fibrillation, and most of the strong inhibitory flavonoids were also found to disaggregate preformed fibrils.

The effect of membranes on the in vitro fibrillation of an amyloidogenic light-chain variable-domain SMA.

Light chain (or AL) amyloidosis is the most common form of systemic amyloidosis, characterized by the pathological deposition of insoluble fibrils of immunoglobulin light-chain fragments in various organs and tissues, especially in the kidney and heart. Both the triggering factors and the mechanisms involved in the abnormal formation of the insoluble fibrillar aggregates from the soluble proteins are poorly understood. For example, although the fibrillar deposits are typically found associated with the extracellular matrix and basement membranes, it is not clear whether fibrils are initially formed intra- or extracellularly, nor it is understood what determines where the deposits will occur; i.e., site tropism. In the present investigation, we studied the interaction of a recombinant amyloidogenic light-chain variable domain, SMA, with lipid vesicles. The nature of the interaction was dependent on the lipid composition and the SMA to lipid ratio. The most pronounced effect was found from vesicles composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphate, which dramatically accelerated fibril growth. Interestingly, spectral probes, such as intrinsic fluorescence and far-UV CD spectroscopy did not show significant conformational changes in the presence of the vesicles. The presence of cholesterol or divalent cations, such as Ca(2+) and Mg(2+), lead to decreased membrane-induced SMA fibrillation. Thus, membranes may have significant effects on light-chain fibrillation and may contribute to the site selectivity observed in AL amyloidosis.