A miR-383-5p signalling hub coordinates the axon regeneration response to inflammation.

Neuroinflammation can positively influence axon regeneration following injury in the central nervous system (CNS). Inflammation promotes the release of neurotrophic molecules and stimulates intrinsic pro-regenerative molecular machinery in neurons, but the detailed mechanisms driving this effect are not fully understood. We evaluated how microRNAs are regulated in retinal neurons in response to intraocular inflammation to identify their potential role in axon regeneration. We found that miR-383-5p is downregulated in retinal ganglion cells in response to zymosan-induced intraocular inflammation. MiR-383-5p downregulation in neurons is sufficient to promote axon growth in vitro, and the intravitreal injection of a miR-383-5p inhibitor into the eye promotes axon regeneration following optic nerve crush. MiR-383-5p directly targets ciliary neurotrophic factor (CNTF) receptor components and miR-383-5p inhibition sensitizes adult retinal neurons to the outgrowth-promoting effects of CNTF. Interestingly, we also demonstrate that CNTF treatment is sufficient to reduce miR-383-5p levels in neurons, constituting a positive-feedback module whereby initial CNTF treatment reduces miR-383-5p levels, which then disinhibits CNTF receptor components to sensitize neurons to the ligand. Additionally, miR-383-5p inhibition de-represses the mitochondrial antioxidant protein peroxiredoxin-3 (PRDX3) which was required for the pro-regenerative effects associated with miR-383-5p loss of function in vitro. We have thus identified a positive feedback mechanism that facilitates neuronal CNTF sensitivity in neurons, and a new molecular signalling module that promotes inflammation-induced axon regeneration.Significance statement Inflammation can both positively and negatively influence the neuronal response to injury. Identifying molecular signalling pathways that mimic pro-regenerative effects of inflammation while bypassing cytotoxic effects is important for our understanding of the precise functions of inflammation in CNS injury and repair. We demonstrate that miR-383-5p is suppressed in neurons in response to inflammatory stimuli and regulates members of the ciliary neurotrophic factor (CNTF) receptor complex, as well as the expression of an antioxidant protein to improve axon regeneration in an optic nerve crush model. These findings identify a new molecular signalling module that promotes axon regeneration and that may bypass detrimental effects of inflammation.

Cardiorespiratory failure induced by inhalation of aerosolized fentanyl in anesthetized rats.

Intravenous rapid injection of fentanyl causes respiratory depression (severe apneas), leading to sudden death, which constitutes the deadliest drug reaction among overdoses of synthetic opioids. Here we asked whether acute inhalation of overdose fentanyl would also result in similar respiratory failure and death. The anesthetized and spontaneously breathing rats with tracheal cannulation were exposed to aerosolized fentanyl at 100 mg/m3 (FNTH) or 30 mg/m3 (FNTL) for 10 min. Minute ventilation (VE), electromyography (EMG) of the internal and external intercostal muscles and thyroarytenoid muscles (EMGII, EMGEI, and EMGTA), heart rate and arterial blood pressure were recorded. During the exposure, FNTH and FNTL immediately triggered bradypnea (40 % reduction, p < 0.05) with TE prolonged and then gradually decreased VE by 40 % (P < 0.05) after a brief VE recovery. The initial TE prolongation (apneas) were characterized by the cessation of EMGEI activity with enhanced tonic discharges of EMGTA and EMGII. After termination of the exposure, the cardiorespiratory responses to FNTL returned to the baseline values 30 min later, while those to FNTH were greatly exacerbated (P < 0.05), leading to ventilatory and cardiac arrest occurred 16.4 ± 4.7 min and 19.3 ± 4.5 min respectively after the onset of FNTH. The ventilatory arrest was featured by cessation of both EMGEI and EMGII and augmentation of tonic EMGTA. Our results suggest that acute exposure to an overdose of fentanyl aerosol leads to death through initially inducing a brief central and upper airway obstructive apnea as well as chest wall rigidity followed by gradual severe hypoventilation, bradycardia and hypotension, and eventual cardiorespiratory arrest in anesthetized rats.

Three new antinociceptive diterpenoids from the fruits of Rhododendron molle.

Investigation of the fruits of Rhododendron molle G. Don led to the isolation of three new grayanane-type diterpenoids, rhodomolleins LIV-LVI (1-3). The structures and absolute configurations of new compounds were fully elucidated by spectroscopic analysis and single-crystal X-ray diffraction, including HRESIMS, 1 D and 2 D NMR data. Compounds 1-3 were evaluated for analgesic activities utilizing an acetic acid-induced writhing test in mice. Compound 1 showed a significant antinociceptive effect with writhe inhibition rates of 72.9% and 100% at doses of 6 mg/kg and 20 mg/kg in mice, respectively. The binding mode of 1 to N-ethylmaleimide-sensitive factor (NSF, PDB: 6IP2) was explored by molecular docking, indicating the presence of hydrogen bond interactions which account for its analgesic activity.

A novel dual-signal output strategy for POCT of CEA based on a smartphone electrochemical aptasensing platform.

A smartphone-based electrochemical aptasensing platform was developed for the point-of-care testing (POCT) of carcinoembryonic antigen (CEA) based on the ferrocene (Fc) and PdPt@PCN-224 dual-signal labeled strategy. The prepared PdPt@PCN-224 nanocomposite showed a strong catalytic property for the reduction of H2O2. Phosphate group-labeled aptamer could capture PdPt@PCN-224 by Zr-O-P bonds to form PdPt@PCN-224-P-Apt. Therefore, a dual signal labeled probe was formed by the hybridization between Fc-DNA and PdPt@PCN-224-P-Apt. The presence of CEA forced PdPt@PCN-224-P-Apt to leave the electrode surface due to the specific affinity, leading to the decrease of the reduction current of H2O2. At the same time, the Fc-DNA strand changed to hairpin structure, which made Fc closer to the electrode and resulted in the increase of the oxidation current of Fc. Thus, CEA can be accurately determined through both signals: the decrease of H2O2 reduction current and the increase of Fc oxidation current, which could avoid the false positive signal. Under the optimal conditions, the prepared aptasensor exhibited a wide linear range from 1 pg·mL-1 to 100 ng·mL-1 and low detection limits of 0.98 pg·mL-1 and 0.27 pg·mL-1 with Fc and PdPt@PCN-224 as signal labels, respectively. The aptasensor developed in this study has successfully demonstrated its capability to detect CEA in real human serum samples. These findings suggest that the proposed sensing platform will hold great potential for clinical tumor diagnosis and monitoring.

Cytotoxic xanthanolide sesquiterpenes from the fruits of Xanthium italicum Moretti.

One previously undescribed xanthanolide sesquiterpene dimer pungiolide P (1), possessing an unprecedented scaffold with a 5/7/5/7/5 ring system skeleton and its intermediate pungiolide Q (2), ten xanthanolide sesquiterpenes (3-12), two eudesmene sesquiterpene derivatives (13-14), one phenylpropionic acid derivative (15), together with eleven known compounds (16-26) were obtained from the fruits of Xanthium italicum Moretti. A possible biosynthetic pathway for pungiolide P (1) was also proposed, which was supported by its bio-synthetic intermediate (2). Compounds 1, 4-5, 18-21, and 25 exhibited cytotoxic activity against a variety of human cancer cell lines. Furthermore, compounds 1, 4-5, could cause blockage of the cell cycle in the G2/M phase and induce apoptosis in H460 cells. Notably, pungiolide P (1) exhibited significantly superior cytotoxicity compared to previously reported compounds, providing valuable insights for natural anti-tumor sources.

Blood Coagulation-Inspired Fibrin Hydrogel for Portable Detection of Thrombin Based on Personal Glucometer.

As one of the biomarkers of coagulation system-related diseases, the detection of thrombin is of practical importance. Thus, this study developed a portable biosensor based on a personal glucometer for rapid detection of thrombin activity. Fibrinogen was used for the detection of thrombin, and the assay principle was inspired by the blood coagulation process, where thrombin hydrolyzes fibrinogen to produce a fibrin hydrogel, and the amount of invertase encapsulated in the fibrin hydrogel fluctuates in accordance with the activity of thrombin in the sample solution. The quantitative assay is conducted by measuring the amount of unencapsulated invertase available to hydrolyze the substrate sucrose, and the signal readout is recorded using a personal glucometer. A linear detection range of 0-0.8 U/mL of thrombin with a limit of detection of 0.04 U/mL was obtained based on the personal glucometer sensing platform. The results of the selectivity and interference experiments showed that the developed personal glucometer sensing platform is highly selective and accurate for thrombin activity. Finally, the reliability of the portable glucometer method for rapid thrombin detection in serum samples was investigated by measuring the recovery rate, which ranged from 92.8% to 107.7%. In summary, the fibrin hydrogel sensing platform proposed in this study offers a portable and versatile means for detecting thrombin using a personal glucometer. This approach not only simplifies the detection process, but also eliminates the need for large instruments and skilled operators, and substantially reduces detection costs.

Natural products in atherosclerosis therapy by targeting PPARs: a review focusing on lipid metabolism and inflammation.

Inflammation and dyslipidemia are critical inducing factors of atherosclerosis. Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors and control the expression of multiple genes that are involved in lipid metabolism and inflammatory responses. However, synthesized PPAR agonists exhibit contrary therapeutic effects and various side effects in atherosclerosis therapy. Natural products are structural diversity and have a good safety. Recent studies find that natural herbs and compounds exhibit attractive therapeutic effects on atherosclerosis by alleviating hyperlipidemia and inflammation through modulation of PPARs. Importantly, the preparation of natural products generally causes significantly lower environmental pollution compared to that of synthesized chemical compounds. Therefore, it is interesting to discover novel PPAR modulator and develop alternative strategies for atherosclerosis therapy based on natural herbs and compounds. This article reviews recent findings, mainly from the year of 2020 to present, about the roles of natural herbs and compounds in regulation of PPARs and their therapeutic effects on atherosclerosis. This article provides alternative strategies and theoretical basis for atherosclerosis therapy using natural herbs and compounds by targeting PPARs, and offers valuable information for researchers that are interested in developing novel PPAR modulators.

Minor Hydroxylated Triterpenoids Produced in Engineered Yeast by the Enzymes OSC and CYP716s from the Plant Enkianthus chinensis and Their Anti-Inflammatory and Hepatoprotective Activities.

Triterpenoids are a type of specialized metabolites that exhibit a wide range of biological activities. However, the availability of some minor triterpenoids in nature is limited, which has hindered our understanding of their pharmacological potential. To overcome this limitation, heterologous biosynthesis of triterpenoids in yeast has emerged as a promising and time-efficient production platform for obtaining these minor compounds. In this study, we analyzed the transcriptomic data of Enkianthus chinensis to identify one oxidosqualene cyclase (EcOSC) gene and four CYP716s. Through heterologous expression of these genes in yeast, nine natural pentacyclic triterpenoids, including three skeleton products (1-3) produced by one multifunctional OSC and six minor oxidation products (4-9) catalyzed by CYP716s, were obtained. Of note, we discovered that CYP716E60 could oxidize ursane-type and oleanane-type triterpenoids to produce 6ß-OH derivatives, marking the first confirmed C-6ß hydroxylation in an ursuane-type triterpenoid. Compound 9 showed moderate inhibitory activity against NO production and dose-dependently reduced IL-1ß and IL-6 production at the transcriptional and protein levels. Compounds 1, 2, 8, and 9 exhibited moderate hepatoprotective activity with the survival rates of HepG2 cells from 61% to 68% at 10 µM.

hP-MSCs attenuate severe acute pancreatitis in mice via inhibiting NLRP3 inflammasome-mediated acinar cell pyroptosis.

BACKGROUND: Severe acute pancreatitis (SAP) is a serious gastrointestinal disease that is facilitated by pancreatic acinar cell death. The protective role of human placental mesenchymal stem cells (hP-MSCs) in SAP has been demonstrated in our previous studies. However, the underlying mechanisms of this therapy remain unclear. Herein, we investigated the regularity of acinar cell pyroptosis during SAP and investigated whether the protective effect of hP-MSCs was associated with the inhibition of acinar cell pyroptosis. METHODS: A mouse model of SAP was established by the retrograde injection of sodium taurocholate (NaTC) solution in the pancreatic duct. For the hP-MSCs group, hP-MSCs were injected via the tail vein and were monitored in vivo. Transmission electron microscopy (TEM) was used to observe the pyroptosis-associated ultramorphology of acinar cells. Immunofluorescence and Western blotting were subsequently used to assess the localization and expression of pyroptosis-associated proteins in acinar cells. Systemic inflammation and local injury-associated parameters were evaluated. RESULTS: Acinar cell pyroptosis was observed during SAP, and the expression of pyroptosis-associated proteins initially increased, peaked at 24 h, and subsequently showed a decreasing trend. hP-MSCs effectively attenuated systemic inflammation and local injury in the SAP model mice. Importantly, hP-MSCs decreased the expression of pyroptosis-associated proteins and the activity of the NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome in acinar cells. CONCLUSIONS: Our study demonstrates the regularity and important role of acinar cell pyroptosis during SAP. hP-MSCs attenuate inflammation and inhibit acinar cell pyroptosis via suppressing NLRP3 inflammasome activation, thereby exerting a protective effect against SAP.

Early-onset Marfan syndrome with aortic dilatation and giant pulmonary artery aneurysm: A case report.

A 30-year-old woman with ankylosing spondylitis was referred to our clinic with abnormal fetal echocardiography findings, including ascending aortic dilatation, giant main pulmonary artery aneurysm, and aortic and pulmonary valve stenosis at 22 weeks of gestation. The full-term male neonate was born by cesarean section and was transferred to the cardiac intensive care unit soon after delivery for respiratory distress with low percutaneous oxygen saturation. Based on cardiovascular and genetic analysis findings, the patient was diagnosed with Marfan syndrome. Surgery was performed; however, the patient died due to cardiac arrest. In conclusion, main pulmonary artery dilatation and aneurysms are uncommon in Marfan syndrome; therefore, presentation with these findings during the fetal life, as in the present case, is likely a sign of severe Marfan syndrome-related cardiac involvement.

Apneic response to fentanyl in adult rats: Role of laryngeal afferents.

Intravenous (systemic) bolus injection of fentanyl (FNT) reportedly induces an immediate vagal-mediated apnea; however, the precise origin of vagal afferents responsible for this apnea remains unknown. We tested whether intralaryngeal (local) application of FNT would also trigger an apnea and whether the apneic response to both local and systemic administration of FNT was laryngeal afferent-mediated. Cardiorespiratory responses to FNT were recorded in anesthetized male adult rats with and without bilateral sectioning of the superior laryngeal nerve (SLNx) or peri-SLN capsaicin treatment (SLNcap) to block local C-fiber signal conduction. Opioid mu-receptor (MOR)-immunoreactivity was detected in laryngeal C- and myelinated neurons. We found that local and systemic administration of FNT elicited an immediate apnea. SLNx, rather than SLNcap, abolished the apneic response to local FNT application though MORs were abundantly expressed in both laryngeal C- and myelinated neurons. Importantly, SLNx failed to affect the apneic response to systemic FNT administration. These results lead to the conclusion that laryngeal afferents' MORs are responsible for the apneic response to local, but not systemic, administration of FNT.

A label-free activatable biosensor for in situ detection of exosomal microRNAs based on DNA-AgNCs and hairpin type nucleic acid probes.

Exosomal microRNA (miRNA) is a potential biomarker for cancer diagnosis, metastasis, and treatment. In situ detection of exosomal miRNA is an attractive option due to its simplicity and high accuracy. However, in situ exosomal miRNA detection has encountered challenges because of the low target abundance of targets and limited probe permeability. Herein, a label-free and activatable biosensor was developed for in situ exosomal miRNA assays by utilizing hairpin-shaped nucleic acid probes and DNA-hosted silver nanoclusters (DNA-AgNCs). The probe is directly internalized into the exosomes, and then hybridized with the target miRNA-21. Subsequently, the DNA-AgNCs are pulled closer to the G-rich sequence, ultimately leading to in situ red fluorescence activation. The biosensor not only can detect exosomal miRNA-21 but also distinguish cancer cells from normal cells. Under optimal reaction conditions, the detection limit (LOD) of exosomal miRNA-21 is 1.53 × 107 particles per mL. Furthermore, DNA-AgNCs are used as label-free signal elements for in situ detection of exosomal miRNAs for the first time, expanding the application of nanomaterials in this field. This strategy does not require tedious RNA extraction steps and expensive instruments, and may develop into a non-invasive diagnostic tool for ovarian cancer.

Characterization and Engineering of Two Highly Paralogous Sesquiterpene Synthases Reveal a Regioselective Reprotonation Switch.

Sesquiterpene synthases (STPSs) catalyze carbocation-driven cyclization reactions that can generate structurally diverse hydrocarbons. The deprotonation-reprotonation process is widely used in STPSs to promote structural diversity, largely attributable to the distinct regio/stereoselective reprotonations. However, the molecular basis for reprotonation regioselectivity remains largely understudied. Herein, we analyzed two highly paralogous STPSs, Artabotrys hexapetalus (-)-cyperene synthase (AhCS) and ishwarane synthase (AhIS), which catalyze reactions that are distinct from the regioselective protonation of germacrene A (GA), resulting in distinct skeletons of 5/5/6 tricyclic (-)-cyperene and 6/6/5/3 tetracyclic ishwarane, respectively. Isotopic labeling experiments demonstrated that these protonations occur at C3 and C6 of GA in AhCS and AhIS, respectively. The cryo-electron microscopy-derived AhCS complex structure provided the structural basis for identifying different key active site residues that may govern their functional disparity. The structure-guided mutagenesis of these residues resulted in successful functional interconversion between AhCS and AhIS, thus targeting the three active site residues [L311-S419-C458]/[M311-V419-A458] that may act as a C3/C6 reprotonation switch for GA. These findings facilitate the rational design or directed evolution of STPSs with structurally diverse skeletons.

A new cytotoxic disaccharide glycoside from the tubers of Arisaema franchetianum.

A new disaccharide glycoside, franchoside A (1), and 17 known compounds were isolated from the tubers of Arisaema franchetianum Engler. The chemical structure of the previously undescribed compound 1 was elucidated on the basis of detailed spectroscopic analyses. Compounds 1, 2, 6, 10, 14 and 18 showed significant cytotoxic activities at varying IC50 values in the range of 4.0-10.6 µM against five cancer cell lines. Compounds 8, 10, 13 and 17 (10 µM) exhibited moderate anti-inflammatory activities by inhibiting the NF-κB signaling pathway and the release of NO from RAW264.7 macrophages induced by lipopolysaccharide (LPS), while compounds 1, 9, 14, 15 and 16 showed weak anti-inflammatory activities.

Identification and characterization of biomarkers associated with endoplasmic reticulum protein processing in cerebral ischemia-reperfusion injury.

Background: Cerebral ischemia (CI), ranking as the second leading global cause of death, is frequently treated by reestablishing blood flow and oxygenation. Paradoxically, this reperfusion can intensify tissue damage, leading to CI-reperfusion injury. This research sought to uncover biomarkers pertaining to protein processing in the endoplasmic reticulum (PER) during CI-reperfusion injury. Methods: We utilized the Gene Expression Omnibus (GEO) dataset GSE163614 to discern differentially expressed genes (DEGs) and single out PER-related DEGs. The functions and pathways of these PER-related DEGs were identified via Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Core genes were pinpointed through protein-protein interaction (PPI) networks. Subsequent to this, genes with diagnostic relevance were distinguished using external validation datasets. A single-sample gene-set enrichment analysis (ssGSEA) was undertaken to pinpoint genes with strong associations to hypoxia and apoptosis, suggesting their potential roles as primary inducers of apoptosis in hypoxic conditions during ischemia-reperfusion injuries. Results: Our study demonstrated that PER-related genes, specifically ADCY5, CAMK2A, PLCB1, NTRK2, and DLG4, were markedly down-regulated in models, exhibiting a robust association with hypoxia and apoptosis. Conclusion: The data indicates that ADCY5, CAMK2A, PLCB1, NTRK2, and DLG4 could be pivotal genes responsible for triggering apoptosis in hypoxic environments during CI-reperfusion injury.

Survey of natural products reported by Asian research groups in 2022.

The new natural products reported in 2022 in peer-reviewed articles in journals with good reputations were reviewed and analyzed. The advances made by Asian research groups in the field of natural products chemistry in 2022 were summarized. Compounds with unique structural features and/or promising bioactivities originating from Asian natural sources were discussed based on their structural classification.

A smartphone-based electrochemical POCT for CEA based on signal amplification of Zr6MOFs.

Carcinoembryonic antigen (CEA) is a biomarker of high expression in cancer cells. Highly sensitive and selective detection of CEA holds significant clinical value in the diagnosis, monitoring and efficacy evaluation of malignant tumors. In this work, a smartphone-based electrochemical point-of-care testing (POCT) platform for the detection of CEA was developed based on a Zr6MOF signal amplification strategy. Ferrocene labeled DNA strands (Fc-DNA) were immobilized on Zr6MOFs to form a Fc-DNA/Zr6MOF signal probe. Double-stranded DNA (dsDNA) formed by complementary DNA (cDNA) and CEA aptamer was assembled on a screen-printed electrode via an Au-S bond. When CEA was added, the aptamer specifically bound with CEA, resulting in the exposure of cDNA. Then, Fc-DNA/Zr6MOF signal probes were introduced on the electrode surface through hybridization between Fc-DNA and cDNA. The detection of CEA was realized by measuring the electrochemical response of Fc. The POCT device was made by connecting a modified electrode with a smartphone through a Sensit Smart USB flash disk. Due to the signal amplification of Zr6MOFs, this POCT platform exhibited high sensitivity, wide linear range, and low detection limit for CEA detection. The developed POCT platform has been used for the detection of CEA in actual human serum samples with satisfactory results.

[Analysis of influence of MR signs on Harris score in ARCO stages 2-4 femoral head necrosis].

OBJECTIVE: To analysis and determine MR signs of Harris score ARCO stages 2-4 in osteonecrosis of femoral head (ONFH). METHODS: Thirty-four patients with ONFH of ARCO stages 2 to 4 who underwent routine MR, T2 mapping, 3D-SPACE sequence examination and Harris score were retrospectively collected from January 2019 to June 2020, and 3 patients were excluded, and 31 patients were finally included, including 23 males and 8 females, aged from 18 to 62 years old with an average of(40.0±10.8) years old. Among them 21 patients with bilateral femoral head necrosis, totally 52 cases, including 17 with ARCO stage 2 patients, 24 ARCO stage 3, and 11 ARCO stage 4. MR imaging signs (femoral head collapse depth, ONFH index, bone marrow edema, hyperplasia, grade and T2 value of cartilage injury, and joint effusion) were scored and measured on the picture archiving and communication system (PACS) workstation, and the cartilage quantitative parameter T2 value was calculated and measured on Siemens postprocessing workstation. Pearson correlation analysis was used to evaluate the correlation between various MR signs and Harris score, and then multiple linear regression analysis was used to examine impact of MR signs on Harris hip score. RESULTS: Femoral head collapse depth(r=-0.563, P=0.000), grade of cartilage injury(r=-0.500, P=0.000), and joint effusion (r=-0.535, P=0.000) were negatively correlated with Harris score by Pearson correlation analysis. Multiple linear regression analysis showed that joint effusion(ß=-6.198, P=0.001) and femoral head collapse depth(ß=-4.085, P=0.014) had a significant negative impact on Harris hip score. CONCLUSION: Femoral head collapse depth and joint effusion both had significant negative relationship with Harris hip score. It is recommended to routinely evaluate femoral head collapse depth and joint effusion quantitatively and gradedly, so as to efficiently and accurately assist clinical diagnosis and treatment.

A label-free electrochemical aptasensor based on Bi-Sb alloy materials for potential POCT of HER-2.

As a prognostic biomarker for breast cancer, human epidermal growth factor receptor 2 (HER-2) is of crucial diagnostic value. Here, a label-free electrochemical aptasensor was established for the ultrasensitive detection of HER-2 using a modified electrode of Bi-Sb alloy materials (Bi-Sb AMs). The performance of the aptasensor was enhanced greatly due to the introduction of Bi-Sb alloy materials (Bi-Sb AMs) with high conductivity. Furthermore, by integrating the aptasensor with the Sensit Smart U-disk electrochemical analyzer, the point-of-care testing (POCT) for HER-2 was realized. Under the optimal experimental parameters, the POCT analyzer showed a wide linear response from 0.01 pg mL-1 to 100 ng mL-1, with a low detection limit (LOD) of 5.96 fg mL-1 for the detection of HER-2. The presented POCT analyzer exhibited good specificity, stability, and reproducibility. Benefiting from the simple operation and rapid testing, the developed analyzer will have potential application in the prognostic diagnosis and treatment of breast cancer.