Design, synthesis, and anti-cancer evaluation of C-14 arylcarbamate derivatives of andrographolide.

In this study, we explored a concise and mild synthetic route to produce novel C-14 arylcarbamate derivatives of andrographolide, a known anti-inflammatory and anticancer natural product. Upon assessing their anti-cancer efficacy against pancreatic ductal adenocarcinoma (PDAC) cells, some derivatives showed stronger cytotoxicity against PANC-1 cells than andrographolide. In addition, we demonstrated one derivative, compound 3m, effectively reduced the expression of oncogenic p53 mutant proteins (p53R273H and p53R248W), proliferation, and migration in PDAC lines, PANC-1 and MIA PaCa-2. Accordingly, the novel derivative holds promise as an anti-cancer agent against pancreatic cancer. In summary, our study broadens the derivative library of andrographolide and develops an arylcarbamate derivative of andrographolide with promising anticancer activity against PDAC.

Assessment of the Performance of Ultrasonography for Detecting Myofascial Trigger Points.

Needle electromyogram (EMG) research has suggested that endplate noise (EPN) is a characteristic of myofascial trigger points (MTrPs). Although several studies have observed MTrPs through ultrasonography, whether they are hyperechoic or hypoechoic in ultrasound images is still controversial. Therefore, this study determined the echogenicity of MTrP ultrasonography. In stage 1, the MTrP of rat masseter muscle was identified through palpation and marked. Needle EMG was performed to detect the presence of EPN. When EPN was detected, ultrasound scans and indwelling needles were used to identify the nodule with a different grayscale relative to that of its surrounding tissue, and the echogenicity of the identified MTrP was determined. In stage 2, these steps were reversed. An ultrasound scan was performed to detect the nodule at the marked site, and an EMG needle was inserted into the nodule to detect EPN. There were 178 recordings in each stage, obtained from 45 rats. The stage 1 results indicate that the MTrPs in ultrasound images were hypoechoic with a 100% sensitivity of assessment. In stage 2, the accuracy and precision of MTrP detection through ultrasonography were 89.9% and 89.2%, respectively. The results indicate that ultrasonography produces highly accurate and precise MTrP detection results.

Noncovalent Association Thermodynamics of Turn-On Fluorescent Probes with Human Serum Albumin: Dual-Concentration Ratio Method.

Efficient quantification of the affinity of a drug and the targeted protein is critical for strategic drug design. Among the various molecules, turn-on fluorescent probes are the most promising signal transducers to reveal the binding strength and site-specificity of designed drugs. However, the conventional method of measuring the binding ability of turn-on fluorescent probes by using the fractional occupancy under the law of mass action is time-consuming and a massive sample is required. Here, we report a new method, called dual-concentration ratio method, for quantifying the binding affinity of fluorescent probes and human serum albumin (HSA). Temperature-dependent fluorescence intensity ratios of a one-to-one complex (L ⋅ HSA) for a turn-on fluorescent probe (L), e. g., ThT (thioflavin T) or DG (dansylglycine), with HSA at two different values of [L]0 /[HSA]0 under the constraint [HSA]0 >[L]0 were collected. The van't Hoff analysis on these association constants further resulted in the thermodynamic properties. Since only two samples at different [L]0 /[HSA]0 are required without the need of [L]0 /[HSA]0 at a wide range, the dual-concentration ratio method is an easy way to greatly reduce the amounts of fluorescent probes and proteins, as well as the acquisition time.

Maternal Th17/Treg Cytokines and Small Extracellular Vesicles in Plasma as Potential Biomarkers for Preeclampsia.

Preeclampsia is one of the most serious pregnancy complications. It may be caused by immunological changes in the early placental microenvironment. The contents of small EVs may serve as biomarkers of pregnancy complications. Evidence suggests that the balance between T helper 17 (Th17) and regulatory T (Treg) cells are critical for preventing preeclampsia. The study recruited 39 pregnant women with preeclampsia and 127 healthy pregnant women. We assessed the levels of both Th17 and Treg cytokines (IL-10, IL-17, IL-21, IL-22, and TGF-ß) in their plasma and small EVs. We found significant differences in the levels of all cytokines in the plasma between the two groups during the second trimester. We also observed significant differences between the two groups in the levels of EV-encapsulated cytokines IL-21, IL-22, and TGF-ß, as well as in total small EVs, during the second trimester. The ROC analysis showed that the classification efficiency (AUC) of TGF-ß in small EVs was 0.81. TGF-ß had the best discriminant ability of all the single EV biomarkers tested, the cross-validation of the accuracy was 0.89. Th17 and Treg cytokines in plasma and small EVs may contribute to maternal immune activation and clarify the potential mechanisms of small EVs and cytokines in preeclampsia.

Outcomes of active cervical therapeutic exercise on dynamic intervertebral foramen changes in neck pain patients with disc herniation.

BACKGROUND: To better understand biomechanical factors that affect intervertebral alignment throughout active therapeutic exercise, it is necessary to determine spinal kinematics when subjects perform spinal exercises. This study aims to investigate the outcomes of active cervical therapeutic exercise on intervertebral foramen changes in neck pain patients with disc herniation. METHODS: Thirty diagnosed C4/5 and/or C5/6 disc-herniated patients receiving an 8-week cervical therapeutic exercise program were followed up with videofluoroscopic images. The dynamic changes in the foramen were computed at different timepoints, including the neutral position, end-range positions in cervical flexion-extension, protrusion-retraction, and lateral flexion movements. RESULTS: The results showed that the active cervical flexion, retraction, and lateral flexion away from the affected side movements increased the area of the patients' intervertebral foramen; while the active extension, protrusion, and lateral flexion toward the affected side reduced the areas of intervertebral foramen before treatment. After the treatment, the active cervical flexion significantly increased the C2/3, C3/4, and C6/7 foramen area by 5.02-8.67% (p = 0.001 ~ 0.029), and the extension exercise significantly reduced the C2/3 and C4/5 area by 5.12-9.18% (p = 0.001 ~ 0.006) compared to the baseline. Active retraction movement significantly increased the foramen area from C2/3 to C6/7 by 3.82-8.66% (p = 0.002 ~ 0.036 with exception of C5/6). Active lateral flexion away from the affected side significantly increased the foramen by 3.71-6.78% (p = 0.007 ~ 0.046 with exception of C6/7). CONCLUSIONS: The 8-week therapeutic exercises including repeated cervical retraction, extension, and lateral flexion movements to the lesion led to significant changes and improvements in intervertebral foramen areas of the patients with disc herniation. TRIAL REGISTRATION: ISRCTN61539024.

Neutralization of IL-10 produced by B cells promotes protective immunity during persistent HCV infection in humanized mice.

Chronic HCV infection can lead to cirrhosis and is associated with increased mortality. Interleukin (IL)-10-producing B cells (B10 cells) are regulatory cells that suppress cellular immune responses. Here, we aimed to determine whether HCV induces B10 cells and assess the roles of the B10 cells during HCV infection. HCV-induced B10 cells were enriched in CD19hi and CD1dhi CD5+ cell populations. HCV predominantly triggered the TLR2-MyD88-NF-κB and AP-1 signaling pathways to drive IL-10 production by B cells. In a humanized murine model of persistent HCV infection, to neutralize IL-10 produced by B10 cells, mice were treated with pcCD19scFv-IL-10R, which contains the genes coding the anti-CD19 single-chain variable fragment (CD19scFv) and the extracellular domain of IL-10 receptor alpha chain (sIL-10Ra). This treatment resulted in significant reduction of B10 cells in spleen and liver, increase of cytotoxic CD8+ T-cell responses against HCV, and low viral loads in infected humanized mice. Our results indicate that targeting B10 cells via neutralization of IL-10 may offer a novel strategy to enhance anti-HCV immunotherapy.

One-pot construction of dispirocyclohexanes via K2CO3-mediated desulfonylative Michael-Michael-aldol (2 + 2 + 2) annulation of sulfonyl 2-pyridyl flavanones and methylketones.

K2CO3-mediated one-pot tandem conversion from sulfonyl 2-pyridyl flavanones to dipyridyl dispirocyclohexanes with contiguous multiple stereogenic centers was accomplished in good yields via an intramolecular desulfonylative ring-contraction of sulfonyl 2-pyridyl flavanones followed by an intermolecular Michael-Michael-aldol (2 + 2 + 2) annulation of the resulting pyridyl aurone intermediate with methyl ketones under facile, open-vessel, inexpensive and environmentally friendly reaction conditions. A plausible mechanism is proposed and discussed herein. This protocol provides a highly effective ring-closure via three carbon-carbon (C-C) bond formation.

Alterations of plasma cytokine biomarkers for identifying age at onset of schizophrenia with neurological soft signs.

Several studies have been suggested that immunity plays a part in neurodevelopment and schizophrenia pathogenesis. Early age of onset in schizophrenia is associated with genetic factors which affect neurodevelopment. This study aims to identify immune abnormalities associated with neurodevelopmental impairments in early-onset schizophrenia (EOS) and adult-onset schizophrenia (AOS) patients. We determined the plasma levels of six cytokines (IL-1ß, IL-4, IL-6, IL-10, IL-12 and TNF-α) in schizophrenia patients and healthy controls. Measurements included neurological soft signs (NSS) to distinguish and subgroup those with neurodevelopmental impairments. The study included 210 schizophrenia patients, which were divided into 84 EOS and 126 AOS patients, as well as 122 healthy controls. We observed significant differences in levels of IL-4, IL-6 and IL-10 between EOS and AOS patients. The results demonstrated the area under ROC curve (AUC) of the IL-4 in EOS and healthy controls was 0.81. Moreover, these results indicated that AUC of the IL-4 and the combination of IL-4, IL-6 and IL-12 in EOS with NSS and healthy controls were 0.91 and 0.95. These cytokines are altered in EOS and schizophrenia patients with neurodevelopmental impairments and demonstrated good classification abilities. These findings manifested that both pro- and anti-inflammatory cytokines are contributed to the clinical and pathophysiological features of schizophrenia. Future works are expected to explore potential genetic effectors and predictors as well as therapeutic directions in personalized medicine for early-onset schizophrenia.

Temperature-Controlled Desulfonylative Condensation of α-Sulfonyl o-Hydroxyacetophenones and 2-Formyl Azaarenes: Synthesis of Azaaryl Aurones and Flavones.

Temperature-controlled intermolecular desulfonylative condensation of α-sulfonyl o-hydroxyacetophenones with 2-formyl azaarenes (pyridines and quinolones) provides azaaryl (pyridyl and quinolyl) aurones and flavones under warming and refluxing toluene reaction conditions via the formation of the intermediate of sulfonyl chroman-4-one. The uses of various solvents are investigated for facile and efficient transformation. A plausible mechanism is proposed.

Intramolecular Benzannulation of 3-Sulfonyl-2-benzylchromen-4-ones: Synthesis of Sulfonyl Dibenzooxabicyclo[3.3.1]nonanes.

In this work, a concise route for the synthesis of sulfonyl dibenzo-oxabicyclo[3.3.1]nonanes by a two-step route is described, including (i) NaBH4/LiCl-mediated reduction of 3-sulfonyl-2-benzylchromen-4-ones and (ii) sequential BF3·OEt2-mediated intramolecular annulation of the resulting 3-sulfonyl-2-benzylchroman-4-ols. A plausible mechanism is proposed and discussed herein. This protocol provides a highly effective stereocontrolled aryl-hydroxyl Friedel-Crafts-type cross-coupling to construct the tetra- or pentacyclic bridged framework. The use of various reaction conditions is investigated for an efficient transformation.

Biodegradation of tetracycline antibiotics in A/O moving-bed biofilm reactor systems.

An anaerobic/aerobic moving-bed biofilm (A/O-MBBR) reactor system was constructed, and the treatment efficiency of aqueous antibiotics in wastewater was investigated. The effects of antibiotics on the microbial communities in the A/O-MBBR were also investigated. Under the optimized reaction conditions, removal of tetracycline antibiotics (TCs) was studied in a series of experiments. When a low concentration of tetracycline (TC) was added to the reactor system, high removal efficiency of conventional pollutants (TC concentration decreased from 10 turn to 2.8 µg L-1) was achieved. When mixed TCs (50 µg L-1) were added to the system, the removal efficiencies of chlortetracycline (CTC), TC and oxytetracycline (OTC) reached 52.03, 41.79, and 38.42%, respectively. TC degradation was decreased to 21.16% when the antibiotic concentration was 500 µg L-1; exposure to this TC concentration destroyed the community structure of the activated sludge bacteria in the reactor. The products of the biodegradation analysis revealed the possible degradation pathways functioning in the experimental A/O-MBBRs.

Regiocontrolled Synthesis of α-Sulfonylmethyl o-Nitrostyrenes via ZnI2-Mediated Sulfonylation and AgNO2/Pd(PPh3)4-Promoted o-Nitration.

We report herein the AgNO2/Pd(PPh3)4-promoted regiocontrolled o-nitration of α-sulfonylmethylstyrenes in MeNO2 with good yields. The o-nitration process provides a series of sulfonyl o-nitrostyrenes. Substituted α-sulfonylmethylstyrenes were synthesized from ZnI2-mediated sulfonylation of substituted α-methylstyrenes and sodium sulfinates (RSO2Na) in MeCN with good to excellent yields. The structures of the key products were confirmed by X-ray crystallography. A plausible mechanism has been proposed herein.

Synthesis of 2-Aryl-3-sulfonylchromans via Knoevenagel Condensation and Reduction Protocol.

In this article, a concise, easy-operational, and high-yield method for the synthesis of 2-aryl-3-sulfonylchromans is described by two-step routes: (i) NH4OAc mediated Knoevenagel condensation of ß-ketosulfones and o-hydroxybenzaldehydes in a cosolvent of toluene and THF at reflux for 10 h, and (ii) NaBH4 promoted regio- and stereocontrolled reduction of the resulting 2-arylchromen-2-ols in a cosolvent of MeOH and THF at rt for 1 h. This protocol provides a highly effective (4+2) annulation via one carbon-oxygen and one carbon-carbon bond formations.

A sensitive immunosorbent bio-barcode assay based on real-time immuno-PCR for detecting 3,4,3',4'-tetrachlorobiphenyl.

A functionalized gold-nanoparticle bio-barcode assay, based on real-time immuno-PCR (IPCR), was designed for the determination of 3,4,3',4'-tetrachlorobiphenyl (PCB77). 15 nm gold nanoparticles were synthesized, and modified with thiol-capped DNA and goat anti-rabbit IgG. The nanoparticle probes were used to replace antibody-DNA conjugate in the IPCR, and were fixed on the PCR tube wall via the immune reaction. Real-time PCR was performed to quantify the DNA signal directly. Under optimized conditions, the new method was used to detect PCB77 with a linearity range from 5 pg L(-1) to 10 ng L(-1), and the limit of detection (LOD) was 1.72 pg L(-1). Real samples of Larimichthys polyactis, collected from the East China Sea, were analyzed. Recovery was from 82 % to 112 %, and the coefficient of variation (CV) was acceptable. The results were compared with GC-ECD, revealing that the method would be acceptable for providing rapid, semi-quantitative, and reliable test results for making environmental decisions.

The shift of segmental contribution ratio in patients with herniated disc during cervical lateral bending.

BACKGROUND: Abnormal intervertebral movements of spine have been reported to be associated with trauma and pathological conditions. The importance of objective spinal motion imaging assessment in the frontal plane was frequently underestimated. The clinical evaluation of the segmental motion contribution could be useful for detecting the motion pattern of individual vertebrae. Therefore the purpose of this study was to investigate the shift of segmental contribution ratio in patients with herniated disc during cervical lateral bending to provide additional insights to cervical biomechanics. METHODS: A total of 92 subjects (46 healthy adult subjects and 46 disc-herniated patients) were enrolled in this case-control study. The motion images during cervical lateral bending movements were digitized using a precise image protocol to analyze the intervertebral motion and contribution. RESULTS: Our results showed that the intervertebral angulation during cervical lateral bending for the C2/3 to C6/7 segments were 7.66°±2.37°, 8.37°±2.11°, 8.91°±3.22°, 7.19°±2.29°, 6.31°±2.11°, respectively for the healthy subjects. For the patients with herniated disc, the intervertebral angulation for the C2/3 to C6/7 segments were 6.87°±1.67°, 7.83°±1.79°, 7.73°±2.71°, 5.13°±2.05°, 4.80°±1.93°, respectively. There were significant angulation and translational differences between healthy subjects and the patients with herniated disc in the C5/6 and C6/7 segments (P=0.001-0.029). The segmental contributions of the individual vertebral segments were further analyzed. There was a significant increase in segmental contribution ratio of C3/4 (P=0.048), while a significant decrease in contribution ratio of C5/6 (P=0.037) was observed in the patients with herniated disc. Our results indicated that the segmental contribution shifted toward the middle cervical spine in the patients with herniated disc. CONCLUSIONS: The segmental contributions of cervical spine during lateral bending movement were first described based on the validated radiographic protocol. The detection of the shift of segmental contribution ratio could be helpful for the diagnosis the motion abnormality resulted from the disc or, facet pathologies, and arthritic changes of cervical spine.

Ent-eudesmane sesquiterpenoids with anti-neuroinflammatory activity from the marine-derived fungus Eutypella sp. F0219.

In this study, twenty-three ent-eudesmane sesquiterpenoids (1-23) including fifteen previously undescribed ones, named eutypelides A-O (1-15) were isolated from the marine-derived fungus Eutypella sp. F0219. Their planar structures and relative configurations were established by HR-ESIMS and extensive 1D and 2D NMR investigations. The absolute configurations of the previously undescribed compounds were determined by single-crystal X-ray diffraction analyses, modified Mosher's method, and ECD calculations. Structurally, eutypelide A (1) is a rare 1,10-seco-ent-eudesmane, whereas 2-15 are typically ent-eudesmanes with 6/6/-fused bicyclic carbon nucleus. The anti-neuroinflammatory activity of all isolated compounds (1-23) was accessed based on their ability to NO production in LPS-stimulated BV2 microglia cells. Compound 16 emerged as the most potent inhibitor. Further mechanistic investigation revealed that compound 16 modulated the inflammatory response by decreasing the protein levels of iNOS and increasing ARG 1 levels, thereby altering the iNOS/ARG 1 ratio and inhibiting macrophage polarization. qRT-PCR analysis showed that compound 16 reversed the LPS-induced upregulation of pro-inflammatory cytokines, including iNOS, TNF-α, IL-6, and IL-1ß, at both the transcriptional and translational levels. These effects were linked to the inhibition of the NF-κB pathway, a key regulator of inflammation. Our findings suggest that compound 16 may be a potential structure basis for developing neuroinflammation-related disease therapeutic agents.

Metaplastic regeneration in the mouse stomach requires a reactive oxygen species pathway.

In pyloric metaplasia, mature gastric chief cells reprogram via an evolutionarily conserved process termed paligenosis to re-enter the cell cycle and become spasmolytic polypeptide-expressing metaplasia (SPEM) cells. Here, we use single-cell RNA sequencing (scRNA-seq) following injury to the murine stomach to analyze mechanisms governing paligenosis at high resolution. Injury causes induced reactive oxygen species (ROS) with coordinated changes in mitochondrial activity and cellular metabolism, requiring the transcriptional mitochondrial regulator Ppargc1a (Pgc1α) and ROS regulator Nf2el2 (Nrf2). Loss of the ROS and mitochondrial control in Ppargc1a-/- mice causes the death of paligenotic cells through ferroptosis. Blocking the cystine transporter SLC7A11(xCT), which is critical in lipid radical detoxification through glutathione peroxidase 4 (GPX4), also increases ferroptosis. Finally, we show that PGC1α-mediated ROS and mitochondrial changes also underlie the paligenosis of pancreatic acinar cells. Altogether, the results detail how metabolic and mitochondrial changes are necessary for injury response, regeneration, and metaplasia in the stomach.

Effect of age on postural performance and control strategies during changes in visual input and dual-tasking stances.

Background: With age, people begin to experience deterioration in standing balance, especially when sensory input is suddenly removed or added. Here, we sought to explore the effects of age on postural performance and postural control strategies. Methods: The convenience sample consisted of 15 young, 10 middle-aged, and 14 elderly healthy adults. They were instructed to stand with their feet together in four randomly administered conditions involving visual input removal/addition and single-/dual-tasking. Dual-tasking involved continuous subtraction by 3s. Results: Postural sway displacement in the two older groups seemed larger than that in the younger group; however, neither the main effect of group (F2, 36 = 1.152, p = .327) nor the group × time interaction effect (F4, 27 = 0.229, p = .922) was significant. Greater stiffness of the lower leg muscles was observed in the vision-addition condition than in the vision-removal condition in only the elderly group (t13 = -2.755, p = .016). The dual-tasking condition resulted in smaller sway displacement (F1, 36 = 7.690, p = .009) and greater muscle stiffness (F1, 36 = 5.495, p = .025). In the vision-removal condition, the increase in muscle stiffness due to dual-tasking was significantly larger in the middle-aged (t9 = -3.736, p = .005) and elderly groups (t13 = -2.512, p = .026). Conclusions: In healthy older individuals, age-related changes were observed in control strategies used to maintain standing balance upon changes in visual input. The dual-task paradigm induced the use of an ankle-stiffening strategy in middle-aged and elderly adults.

Single-cell sequencing of ascites fluid illustrates heterogeneity and therapy-induced evolution during gastric cancer peritoneal metastasis.

Peritoneal metastasis is the leading cause of death for gastrointestinal cancers. The native and therapy-induced ascites ecosystems are not fully understood. Here, we characterize single-cell transcriptomes of 191,987 ascites cancer/immune cells from 35 patients with/without gastric cancer peritoneal metastasis (GCPM). During GCPM progression, an increase is seen of monocyte-like dendritic cells (DCs) that are pro-angiogenic with reduced antigen-presenting capacity and correlate with poor gastric cancer (GC) prognosis. We also describe the evolution of monocyte-like DCs and regulatory and proliferative T cells following therapy. Moreover, we track GC evolution, identifying high-plasticity GC clusters that exhibit a propensity to shift to a high-proliferative phenotype. Transitions occur via the recently described, autophagy-dependent plasticity program, paligenosis. Two autophagy-related genes (MARCKS and TXNIP) mark high-plasticity GC with poorer prognosis, and autophagy inhibitors induce apoptosis in patient-derived organoids. Our findings provide insights into the developmental trajectories of cancer/immune cells underlying GCPM progression and therapy resistance.

Role of Integrin αvß3 in Doxycycline-Induced Anti-Proliferation in Breast Cancer Cells.

Doxycycline, an antibiotic, displays the inhibition of different signal transduction pathways, such as anti-inflammation and anti-proliferation, in different types of cancers. However, the anti-cancer mechanisms of doxycycline via integrin αvß3 are incompletely understood. Integrin αvß3 is a cell-surface anchor protein. It is the target for estrogen, androgen, and thyroid hormone and plays a pivotal role in the proliferation, migration, and angiogenic process in cancer cells. In our previous study, thyroxine hormones can interact with integrin αvß3 to activate the extracellular signal-regulated kinase 1/2 (ERK1/2), and upregulate programmed death-ligand 1 (PD-L1) expression. In the current study, we investigated the inhibitory effects of doxycycline on proliferation in two breast cancer cell lines, MCF-7 and MDA-MB-231 cells. Doxycycline induces concentration-dependent anti-proliferation in both breast cancer cell lines. It regulates gene expressions involved in proliferation, pro-apoptosis, and angiogenesis. Doxycycline suppresses cell cyclin D1 (CCND1) and c-Myc which play crucial roles in proliferation. It also inhibits PD-L1 gene expression. Our findings show that modulation on integrin αvß3 binding activities changed both thyroxine- and doxycycline-induced signal transductions by an integrin αvß3 inhibitor (HSDVHK-NH2). Doxycycline activates phosphorylation of focal adhesion kinase (FAK), a downstream of integrin, but inhibits the ERK1/2 phosphorylation. Regardless, doxycycline-induced FAK phosphorylation is blocked by HSDVHK-NH2. In addition, the specific mechanism of action associated with pERK1/2 inhibition via integrin αvß3 is unknown for doxycycline treatment. On the other hand, our findings indicated that inhibiting ERK1/2 activation leads to suppression of PD-L1 expression by doxycycline treatment. Furthermore, doxycycline-induced gene expressions are disturbed by a specific integrin αvß3 inhibitor (HSDVHK-NH2) or a mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinases (ERK) kinase (MAPK/ERK, MEK) inhibitor (PD98059). The results imply that doxycycline may interact with integrin αvß3 and inhibits ERK1/2 activation, thereby regulating cell proliferation and downregulating PD-L1 gene expression in estrogen receptor (ER)-negative breast cancer MDA-MB-231 cells.