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The ability to predict or detect colorectal cancer (CRC) recurrence early after surgery enables physicians to apply appropriate treatment plans and different follow-up strategies to improve patient survival. Overall, 30-50% of CRC patients experience cancer recurrence after radical surgery, but current surveillance tools have limitations in the precise and early detection of cancer recurrence. Circulating tumor cells (CTCs) are cancer cells that detach from the primary tumor and enter the bloodstream. These can provide real-time information on disease status. CTCs might become novel markers for predicting CRC recurrence and, more importantly, for making decisions about additional adjuvant chemotherapy. In this review, the clinical application of CTCs as a therapeutic marker for stage II CRC is described. It then discusses the utility of CTCs for monitoring cancer recurrence in advanced rectal cancer patients who undergo neoadjuvant chemoradiotherapy. Finally, it discusses the roles of CTC subtypes and CTCs combined with clinicopathological factors in establishing a multimarker model for predicting CRC recurrence.
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Optically induced dielectrophoresis (ODEP)-based microparticle sorting and separation is regarded as promising. However, current methods normally lack the downstream process for the transportation and collection of separated microparticles, which could limit its applications. To address this issue, an ODEP microfluidic chip encompassing three microchannels that join only at the central part of the microchannels (i.e., the working zone) was designed. During operation, three laminar flows were generated in the zone, where two dynamic light bar arrays were designed to sort and separate PS (polystyrene) microbeads of different sizes in a continuous manner. The separated PS microbeads were then continuously transported in laminar flows in a partition manner for the final collection. The results revealed that the method was capable of sorting and separating PS microbeads in a high-purity manner (e.g., the microbead purity values were 89.9 ± 3.7, 88.0 ± 2.5, and 92.8 ± 6.5% for the 5.8, 10.8, and 15.8 µm microbeads harvested, respectively). Overall, this study demonstrated the use of laminar flow and ODEP to achieve size-based sorting, separation, and collection of microparticles in a continuous and high-performance manner. Apart from the demonstration, this method can also be utilized for size-based sorting and the separation of other biological or nonbiological microparticles.
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Electroforesis , Técnicas Analíticas Microfluídicas , Microesferas , Tamaño de la Partícula , Poliestirenos , MicrofluídicaRESUMEN
The analysis of circulating tumor cells (CTCs) at the molecular level holds great promise for several clinical applications. For this goal, the harvest of high-purity, size-sorted CTCs with different subtypes from a blood sample are important. For this purpose, a two-step CTC isolation protocol was proposed, by which the immunomagnetic beads-based cell separation was first utilized to remove the majority of blood cells. After that, an optically induced dielectrophoresis (ODEP) microfluidic system was developed to (1) purify the CTCs from the remaining magnetic microbeads-bound blood cells and to (2) sort and separate the CTCs with different sizes. In this study, the ODEP microfluidic system was designed and fabricated. Moreover, its optimum operation conditions and performance were explored. The results exhibited that the presented technique was able to purify and sort the cancer cells with two different sizes from a tested cell suspension in a high-purity (93.5% and 90.1% for the OECM 1 and HA22T cancer cells, respectively) manner. Overall, this study presented a technique for the purification and sorting of cancer cells with different sizes. Apart from this application, the technique is also useful for other applications in which the high-purity and label-free purification and sorting of cells with different sizes is required.
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For the rapid detection of bacteria in a blood sample, nucleic acid amplification-based assays are believed to be promising. Nevertheless, the nucleic acids released from the dead blood cells or bacteria could affect the assay performance. This highlights the importance of the isolation of live bacteria from blood samples. To address this issue, this study proposes a two-step process. First, a blood sample was treated with the immuno-magnetic microbeads-based separation to remove the majority of blood cells. Second, an optically induced dielectrophoresis (ODEP) microfluidic system with an integrated dynamic circular light image array was utilized to further isolate and purify the live bacteria from the remaining blood cells based on their size difference. In this work, the ODEP microfluidic system was developed. Its performance for the isolation and purification of bacteria was evaluated. The results revealed that the method was able to harvest the live bacteria in a high purity (90.5~99.2%) manner. Overall, the proposed method was proven to be capable of isolating and purifying high-purity live bacteria without causing damage to the co-existing cells. This technical feature was found to be valuable for the subsequent nucleic-acid-based bacteria detection, in which the interferences caused by the nontarget nucleic acids could be eliminated.
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Técnicas Analíticas Microfluídicas , Ácidos Nucleicos , Microfluídica , BacteriasRESUMEN
Analysis of circulating tumor cells (CTCs) holds promise to diagnose cancer or monitor its development. Among the methods, counting CTC numbers in blood samples could be the simplest way to implement it. Nevertheless, its clinical utility has not yet been fully accepted. The reasons could be due to the rarity and heterogeneity of CTCs in blood samples that could lead to misleading results from assays only based on single CTC counts. To address this issue, a feasible direction is to combine the CTC counts with other clinical data for analysis. Recent studies have demonstrated the use of this new strategy for early detection and prognosis evaluation of cancers, or even for the distinguishment of cancers with different stages. Overall, this approach could pave a new path to improve the technical problems in the clinical applications of CTC counting techniques. In this review, the information relevant to CTCs, including their characteristics, clinical use of CTC counting, and technologies for CTC enrichment, were first introduced. This was followed by discussing the challenges and new perspectives of CTC counting techniques for clinical applications. Finally, the advantages and the recent progress in combining CTC counts with other clinical parameters for clinical applications have been discussed.
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BACKGROUND: Tumor staging is crucial for melanoma, of which acral melanoma is the predominant subtype in Asians. 18 F-fluoro-2-deoxyglucose (FDG) positron emission tomography (PET) and 18 F-FDG-PET/computed tomography ( 18 F-FDG-PET/CT) serve as noninvasive imaging tools for tumor staging. However, the literature is scarce on the diagnostic value of PET for acral melanoma. METHODS: From January 1, 2006 to November 30, 2022, a total of 352 patients were diagnosed with melanoma at our hospital. Of them, 90 were diagnosed with cutaneous melanoma and underwent preoperative PET/CT for staging and sentinel lymph node biopsy or complete lymph node dissection. Staging of PET/CT was confirmed by histopathology or following imaging. The lymph node biopsy, distant metastasis status, and PET/CT imaging results were analyzed. RESULTS: Of all the 90 patients with cutaneous melanoma, 72 of them were diagnosed as acral melanoma (80.0%). Compared with the histopathologic results, the lymph nodes were true-positive, true-negative, false-positive, and false-negative in 12, 54, 7, and 17 cases, respectively. The sensitivity of PET/CT for local lymph nodes was 41.4% (95% CI, 23.5%-61.1%), whereas its specificity was 88.5% (95% CI, 77.8%-95.3%). As for the detection of distal metastasis, the PET results were true-positive, true-negative, false-positive, and false-negative in 6, 65, 15, and 4 cases, respectively. The sensitivity of PET for distal metastasis detection was 60.0% (95% CI, 26.2%-87.8%), whereas its specificity was 81.3% (95% CI, 71.0%-89.1%). CONCLUSION: Although noninvasive, PET/CT has relatively low sensitivity in regional lymph node evaluations, and fair sensitivity in distal metastasis detection in Asian patients with acral melanoma. Thus, PET/CT may be more useful in patients with clinically palpable nodes or more advanced disease stages.
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Melanoma , Neoplasias Cutáneas , Humanos , Tomografía Computarizada por Tomografía de Emisión de Positrones , Fluorodesoxiglucosa F18 , Radiofármacos , Metástasis Linfática/patología , Sensibilidad y Especificidad , Biopsia del Ganglio Linfático Centinela , Tomografía de Emisión de Positrones/métodos , Ganglios Linfáticos/patología , Estadificación de Neoplasias , Melanoma Cutáneo MalignoRESUMEN
BACKGROUND: Sentinel lymph node (SLN) status is the predominant prognostic factor in patients diagnosed with clinically localized melanoma. The significance of completion lymph node dissection in patients with SLN metastasis is debatable. Not many studies have been conducted on acrallentiginous melanoma (ALM). This study aimed to characterize the prognostic factors of nodal positive ALM and confirm whether ALM patients can undergo the same treatment strategy as non-ALM patients in the Asian population. METHODS: This is a retrospective review of patients who underwent surgery for cutaneous melanoma (CM) at Taipei Veterans General Hospital between January 1993 and December 2019. We investigated the risk factors for lymph node status. The association between clinicopathological factors and lymph node status of ALM and non-ALM patients was analyzed. Outcomes of completion lymph node dissection (CLND) performed following sentinel lymph node biopsy (SLNB) in the CM and ALM groups were compared. RESULTS: A total of 197 patients were included in this study. ALM was the most common histological subtype, accounting for 66.5% of all the cases. Patients in the CM and ALM subgroups with metastatic SLN ( p = 0.012) or lymph nodes ( p < 0.001 and p = 0.001) exhibited higher mortality rate. Multivariate analysis showed that patients with clinical presentation of T4 category tumor ( p = 0.012) and lymphovascular invasion ( p = 0.012) had a significantly higher risk of positive lymph nodes. The overall survival of patients with lymph nodes metastasis was not associated with the performance of CLND. CONCLUSION: Patients in the CM or ALM subgroups with metastatic SLNs or lymph nodes exhibited significantly poorer overall survival. Advanced Breslow thickness and lymphovascular invasion were independent predictive factors for CM and ALM patients with positive lymph node status. There was no significant difference in survival between CM and ALM patients following SLNB, regardless of CLND being performed.
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Melanoma , Neoplasias Cutáneas , Humanos , Melanoma/cirugía , Neoplasias Cutáneas/cirugía , Biopsia del Ganglio Linfático Centinela , Metástasis Linfática/patología , Escisión del Ganglio Linfático , Ganglios Linfáticos/patología , Estudios Retrospectivos , Pronóstico , Melanoma Cutáneo MalignoRESUMEN
Magnetic beads improve biosensing performance by means of their small volume and controllability by magnetic force. In this study, a new technique composed of optically induced dielectrodphoresis (ODEP) manipulation and image processing was used to enhance the signal-to-noise ratio of the fluorescence for stained magnetic beads. According to natural advantages of size-dependent particle isolation by ODEP manipulation, biomarkers in clinical samples can be easily separated by different sizes of magnetic beads with corresponding captured antibodies, and rapidly distinguished by separated location of immunofluorescence. To verify the feasibility of the concept, magnetic beads with three different diameters, including 21.8, 8.7, and 4.2 µm, were easily separated and collected into specific patterns in the defined target zone treated as three dynamic transducer elements to evaluate fluorescence results. In magnetic beads with diameter of 4.2 µm, the lowest signal-to-noise ratio between stained and nonstained magnetic beads was 3.5. With the help of ODEP accumulation and detection threshold setting of 32, the signal-to-noise ratio was increased to 77.4, which makes this method more reliable. With the further optimization of specific antibodies immobilized on different-size magnetic beads in the future, this platform can be a potential candidate for a high-efficiency sensor array in clinical applications.
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Técnicas Biosensibles , Separación Inmunomagnética , Técnicas Biosensibles/métodos , Magnetismo , Relación Señal-Ruido , TransductoresRESUMEN
Optically induced dielectrophoresis (ODEP) is effective for cell manipulation. However, its utilization has been limited by the requirement of solution with low conductivity. This issue has been ignored in ODEP-relevant studies. To address this issue, this study aims to investigate to what extent the cell viability and performance of ODEP-based cell manipulation are affected by low conductivity conditions. Additionally, this study aims to modify sucrose solutions to reduce the impacts caused by low-conductivity solutions. Results revealed the use of sucrose solution in ODEP operation could significantly reduce the viability of the manipulated cells by 9.1 and 38.5% after 2- and 4-h incubation, respectively. Prolonged operation time (e.g., 4 h) in sucrose solution could lead to significantly inferior performance of cell manipulation, including 47.2% reduction of ODEP manipulation velocity and 44.4% loss of the cells manipulatable by ODEP. The key finding of this study is that the use of bovine serum albumin (BSA)-supplemented sucrose solution (conductivity: 25-50 µS cm-1) might significantly increase the cell viability by 10.9-14.8% compared with that in sucrose solution after 4 h incubation. Moreover, the ODEP manipulation velocity of cells in the BSA-supplemented sucrose solution (conductivity: 25 µS cm-1) was comparable to that in sucrose solution during 4-h incubation. More importantly, compared with sucrose solution, the use of BSA-supplemented sucrose solution (conductivity: 25-50 µS cm-1) contributed high percentage (80.4-93.5%) of the cells manipulatable by ODEP during 4-h incubation. Overall, this study has provided some fundamental information relevant to the improvement of background solutions for ODEP-based cell manipulation.
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BACKGROUND: The goal of this study was to investigate the prognostic factors for locoregional recurrence and metastasis in patients with cutaneous melanoma (CM) who underwent surgery, especially in the acral lentiginous melanoma (ALM) subtype. METHODS: This study was a retrospective review of patients who underwent surgery for CM at Taipei Veterans General Hospital between 2000 and 2018. We investigated the risk factors for locoregional and distant metastases. The association between clinicopathological factors and locoregional recurrence and distant metastasis of the CM and ALM subtypes was analyzed. In addition, the outcomes between the ALM and non-ALM groups were compared. RESULTS: A total of 161 patients were included in the analysis. The most common histological subtype was ALM. The overall locoregional recurrence rate of CM was 13.0% and the distant metastasis rate was 42.9%, whereas that of the ALM subtype was 12.5% and 45.5%, respectively. In patients with CM, male sex, tumor with lymphovascular invasion, and positive lymph node status were the prognostic factors for both locoregional recurrence and distant metastasis. Among the patients with ALM, positive lymph node status was significantly associated with both locoregional recurrence and distant metastasis. CONCLUSION: In this cohort, factors influencing locoregional recurrence and distant metastasis were similar between the ALM and non-ALM groups. The above-recommended surgical margin did not show any benefit in either the CM or the ALM subtype. ALM can be handled using the same surgical strategy as CM in the Asian population.
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Melanoma , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/etiología , Neoplasias Cutáneas , Anciano , Femenino , Humanos , Masculino , Melanoma/cirugía , Persona de Mediana Edad , Periodo Posoperatorio , Estudios Retrospectivos , Neoplasias Cutáneas/cirugía , Taiwán , Centros de Atención Terciaria , Melanoma Cutáneo MalignoRESUMEN
Conventional circulating tumor cell (CTC) enumeration could ignore the CTCs more relevant to cancer metastasis. Thus, negative selection CTC enumeration was proposed, by which information on two cellular biomarkers (numbers of CTCs and CD45neg EpCAMneg cells) can be obtained. By combining this approach with the conventional biomarker carcinoembryonic antigen (CEA), this study aimed to explore whether any combination of these biomarkers could improve the predictive performance for colorectal cancer (CRC) or its status. In this work, these two cell populations in healthy donors and CRC patients were quantified. Results revealed that enumeration of these two cell populations was able to discriminate healthy donors from CRC patients, even patients with non-advanced CRC. Moreover, the combination of the two cell populations showed improved performance (AUROC: 0.893) for CRC prediction over the use of only one population. Compared with CEA alone, the combination of the three biomarkers increased the performance (AUROC) for advanced CRC prediction from 0.643 to 0.727. Compared with that of CEA alone for metastatic CRC prediction, the AUROC was increased from 0.780 to 0.837 when the CTC count was included. Overall, this study demonstrated that the combination of these two cellular biomarkers with CEA improved the predictive performance for CRC and its status.
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To develop a lab on a chip (LOC) integrated with both sensor and actuator functions, a novel two-in-one system based on optical-driven manipulation and sensing in a microfluidics setup based on a hydrogenated amorphous silicon (a-Si:H) layer on an indium tin oxide/glass is first realized. A high-intensity discharge xenon lamp functioned as the light source, a chopper functioned as the modulated illumination for a certain frequency, and a self-designed optical path projected on the digital micromirror device controlled by the digital light processing module was established as the illumination input signal with the ability of dynamic movement of projected patterns. For light-addressable potentiometric sensor (LAPS) operation, alternating current (AC)-modulated illumination with a frequency of 800 Hz can be generated by the rotation speed of the chopper for photocurrent vs bias voltage characterization. The pH sensitivity, drift coefficient, and hysteresis width of the Si3N4 LAPS are 52.8 mV/pH, -3.2 mV/h, and 10.5 mV, respectively, which are comparable to the results from the conventional setup. With an identical two-in-one system, direct current illumination without chopper rotation and an AC bias voltage can be provided to an a-Si:H chip with a manipulation speed of 20 µm/s for magnetic beads with a diameter of 1 µm. The collection of magnetic beads by this light-actuated AC electroosmosis (LACE) operation at a frequency of 10 kHz can be easily realized. A fully customized design of an illumination path with less decay can be suggested to obtain a high efficiency of manipulation and a high signal-to-noise ratio of sensing. With this proposed setup, a potential LOC system based on LACE and LAPS is verified with the integration of a sensor and an actuator in a microfluidics setup for future point-of-care testing applications.
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The transcription regulatory network inside a eukaryotic cell is defined by the combinatorial actions of transcription factors (TFs). However, TF binding studies in plants are too few in number to produce a general picture of this complex network. In this study, we use large-scale ChIP-seq to reconstruct it in the maize leaf, and train machine-learning models to predict TF binding and co-localization. The resulting network covers 77% of the expressed genes, and shows a scale-free topology and functional modularity like a real-world network. TF binding sequence preferences are conserved within family, while co-binding could be key for their binding specificity. Cross-species comparison shows that core network nodes at the top of the transmission of information being more conserved than those at the bottom. This study reveals the complex and redundant nature of the plant transcription regulatory network, and sheds light on its architecture, organizing principle and evolutionary trajectory.
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Redes Reguladoras de Genes , Hojas de la Planta/genética , Factores de Transcripción/genética , Zea mays/genética , Secuenciación de Inmunoprecipitación de Cromatina , Biología Computacional/métodos , Aprendizaje Automático , Proteínas de Plantas/genética , Poaceae/genética , Factores de Transcripción/metabolismoRESUMEN
Negative selection-based circulating tumor cell (CTC) isolation is able to harvest viable, label-free, and clinically meaningful CTCs from the cancer patients' blood. Nevertheless, its main shortcoming is its inability to isolate high-purity CTCs, restricting subsequent CTC-related analysis. To address this issue, this study proposed a two-step optically-induced dielectrophoresis (ODEP) cell manipulation to process the cell sample harvested by negative selection-/immunomagnetic microbeads-based CTC isolation. The working mechanism is that the ODEP force acting on the cells with and without magnetic microbeads binding is different. Accordingly, the use of ODEP cell manipulation in a microfluidic system was designed to first separate and then isolate the cancer cells from other magnetic microbead-bound cells. Immunofluorescent microscopic observation and ODEP cell manipulation were then performed to refine the purity of the cancer cells. In this study, the optimum operating conditions for effective cell isolation were determined experimentally. The results revealed that the presented method was able to further refine the purity of cancer cell in the sample obtained after negative selection-based CTC isolation with high cell purity (81.6~86.1%). Overall, this study proposed the combination of immunomagnetic bead-based cell isolation and ODEP cell manipulation for the negative selection-based isolation of CTCs.
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Cell manipulation using optically induced dielectrophoresis (ODEP) in microfluidic systems has attracted the interest of scientists due to its simplicity. Although this technique has been successfully demonstrated for various applications, one fundamental issue has to be addressed-Whether, the ODEP field affects the native properties of cells. To address this issue, we explored the effect of ODEP electrical conditions on cellular properties. Within the experimental conditions tested, the ODEP-based cell manipulation with the largest velocity occurred at 10 Vpp and 1 MHz, for the two cancer cell types explored. Under this operating condition, however, the cell viability of cancer cells was significantly affected (e.g., 70.5 ± 10.0% and 50.6 ± 9.2% reduction for the PC-3 and SK-BR-3 cancer cells, respectively). Conversely, the exposure of cancer cells to the ODEP electrical conditions of 7-10 Vpp and 3-5 MHz did not significantly alter the cell viability, cell metabolic activity, and the EpCAM, VIM, and ABCC1 gene expression of cancer cells. Overall, this study fundamentally investigated the effect of ODEP electrical conditions on the cellular properties of cancer cells. The information obtained is crucially important for the utilization of ODEP-based cell manipulation in a microscale system for various applications.
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Técnicas Biosensibles , Técnicas Analíticas Microfluídicas , Imagen Óptica , Línea Celular Tumoral , Humanos , Células PC-3RESUMEN
The spatial organization of the genome plays an important role in the regulation of gene expression. However, the core structural features of animal genomes, such as topologically associated domains (TADs) and chromatin loops, are not prominent in the extremely compact Arabidopsis genome. In this study, we examine the chromatin architecture, as well as their DNA methylation, histone modifications, accessible chromatin, and gene expression, of maize, tomato, sorghum, foxtail millet, and rice with genome sizes ranging from 0.4 to 2.4 Gb. We found that these plant genomes can be divided into mammalian-like A/B compartments. At higher resolution, the chromosomes of these plants can be further partitioned to local A/B compartments that reflect their euchromatin, heterochromatin, and polycomb status. Chromatins in all these plants are organized into domains that are not conserved across species. They show similarity to the Drosophila compartment domains, and are clustered into active, polycomb, repressive, and intermediate types based on their transcriptional activities and epigenetic signatures, with domain border overlaps with the local A/B compartment junctions. In the large maize and tomato genomes, we observed extensive chromatin loops. However, unlike the mammalian chromatin loops that are enriched at the TAD border, plant chromatin loops are often formed between gene islands outside the repressive domains and are closely associated with active compartments. Our study indicates that plants have complex and unique 3D chromatin architectures, which require further study to elucidate their biological functions.
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Ensamble y Desensamble de Cromatina/genética , Cromatina/genética , Genoma de Planta/genética , Heterocromatina/genética , Histonas/metabolismoRESUMEN
BACKGROUND/PURPOSE: House dust mite (HDM) is well known as one of the major indoor allergens that trigger allergic inflammation, especially asthma, and accounts for 85% of all cases. So far, asthma has been thought of as a condition of imbalance between T helper (Th)1 and Th2. Fungal immunomodulatory protein-Flammulina velutipes (FIP-fve) has been seemingly demonstrated to modulate the response to Th1 cytokine production. The aim of this study was to investigate if the oral administration of FIP-fve can inhibit HDM-induced asthma inflammation in the mouse model. METHODS: We divided the mice (female BALB/c, 4-6 weeks) into four groups: the prevention group, which consisted of mice sensitized by HDM (intraperitoneally on Day 1, Day 7, and Day 14, and intranasally on Day 14, Day 17, Day 21, Day 24, and Day 27) fed with FIP-fve from Day 1 to Day 14; the treatment group, which comprised mice that received treatment from Day 14 to Day 28; the positive control (PC, sensitized by HDM fed without FIP-fve) group; and the negative control group (NC, nonsensitized). Airway hyperresponsiveness induced by methacholine challenge was determined using whole-body barometric plethysmography. In addition, cytokines were analyzed from bronchoalveolar lavage fluid and serum. Histopathological studies and Liu's staining method in mice lungs were also performed. RESULTS: The results showed that both pre- and posttreated FIP-fve groups had significantly reduced airway hyperresponsiveness compared with the PC group after methacholine challenge. In addition, a significantly decreased level of HDM-specific immunoglobulin E in serum and decreased production of Th2 cytokines in bronchoalveolar lavage fluid and serum were observed in these two FIP-fve fed groups. Moreover, more decreased amounts of infiltrating inflammatory cells were present in the lungs of FIP-fve fed groups than those of the PC group. CONCLUSION: Oral FIP-fve had an anti-inflammatory effect on the acute phase of the airway inflammatory process induced by HDM in the mouse model and might have a potentially therapeutic role for allergic airway diseases.
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Asma/tratamiento farmacológico , Flammulina/química , Proteínas Fúngicas/administración & dosificación , Factores Inmunológicos/administración & dosificación , Administración Oral , Animales , Modelos Animales de Enfermedad , Femenino , Proteínas Fúngicas/aislamiento & purificación , Histocitoquímica , Factores Inmunológicos/aislamiento & purificación , Ratones Endogámicos BALB C , Pletismografía , Pyroglyphidae/inmunología , Resultado del TratamientoRESUMEN
Neonatal adrenal hemorrhage (NAH) is rare and is found in only 0.2% of newborns. Scrotal hematoma (SH) in newborns is also rare. NAH associated with SH is extremely rare, with only 29 cases reported in the literature. Herein, we report a baby boy who presented with SH; after ultrasonography examinations, the diagnosis of NAH associated with SH was made. He received conservative treatment only. From our experience and that of others, appropriate integration of clinical information, physical examination and the results of abdominal and scrotal ultrasonography can achieve the accurate diagnosis of NAH associated with SH. This association allows conservative treatment that avoids unnecessary surgical exploration.