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Liang-Jian Hong, Ai-Jun Chen, Feng-Zeng Li, Ke-Jun Chen, Sheng Fang Department of Dermatology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, People’s Republic of ChinaCorrespondence: Sheng FangDepartment of Dermatology, The First Affiliated Hospital of Chongqing Medical University, 1 Youyi Road, Yuzhong District, Chongqing, People’s Republic of ChinaEmail fangshengderm@163.comObjective: To explore the molecular mechanism of 17-AAG in the treatment of systemic lupus erythematosus (SLE), and the effects of the heat shock protein 90 (HSP90) inhibitor 17-AAG on the activation and proliferation of lymphocytes and the AKT/GSK3β signaling pathway in MRL/lpr mice were detected.Methods: MRL/lpr mice were randomly divided into the control group and the experimental group. The experimental group was injected intraperitoneally with 17-AAG, and T lymphocytes were separated by magnetic beads. Lymphocyte proliferation was detected by MTT and flow cytometry (FCM), and the expression of the HSP90 protein and PI3K/AKT signaling pathway-related proteins was detected by Western blotting. Renal histopathology and immune complex deposition were also observed in both groups.Results: Immune complex deposition and inflammation decreased in kidneys from MRL/lpr mice in the experimental group. HSP90 protein expression, T lymphocyte proliferation and phosphorylated AKT and GSK3β levels also decreased in the experimental group.Conclusion: 17-AAG can inhibit the activation and proliferation of T lymphocytes and downregulate the AKT/GSK3β signaling pathway, which may be relevant for the treatment of SLE.Keywords: 17-AAG, HSP90, AKT, lupus, activation

We aimed to determine the incidence, treatment regimen, and treatment outcomes (including progression-free survival and overall survival) of gynecologic carcinosarcoma, a rare, aggressive, and understudied gynecologic malignancy. This retrospective review included all patients with gynecologic cancers diagnosed and treated at a single tertiary care comprehensive cancer center between January 2012 and May 2021. A total of 2116 patients were eligible for review, of which 84 cases were identified as carcinosarcoma: 66 were uterine (5.2% of uterine cancers), 17 were ovarian (3.6% of ovarian cancers), 1 was cervical (0.28% of cervical cancers), and 1 was untyped. Of the patients, 76.2% presented advanced-stage disease (stage III/IV) at the time of diagnosis. Minority patients were more likely to present with stage III/IV ( p < 0.0001). The majority of patients underwent surgical resection followed by systemic chemotherapy with carboplatin and paclitaxel. The median PFS was 7.5 months. Of the patients, 55% were alive 1 year after diagnosis, and 45% were alive at 5 years. In the studied population, minorities were more likely to present with more advanced disease. The rate of gynecologic carcinosarcomas was consistent with historical reports.

Objective: Women make up a majority of the gynecologic oncology workforce. Increasing the numbers of women in leadership has been proposed as a path towards professional gender equity. This study examined whether leadership gender and departmental infrastructure impact the work environment for women gynecologic oncologists., Methods: Members of a 472-member private Facebook group "Women of Gynecologic Oncology" (WGO) who self-identified as women gynecologic oncologists provided demographics, practice infrastructure, personal experience with workplace bullying, gender discrimination, microaggressions using a REDcap survey platform., Results: Of 250 (53%) respondents to this survey, most were younger than age 50 years (93.6%); White (82.2%) and non-Hispanic (94.3%); married (84.7%); and parenting (75.2%). Practice environments included academic (n=152, 61.0%), hospital employed (n=57, 22.9%), and private practice (n=31, 12.4%), and 89.9% supervised trainees. A significant percent of respondents had experienced bullying (52.8%), gender discrimination (57%) and microaggressions (83%). Age, race, ethnicity, practice setting, or mentorship were not statistically significantly associated with these experiences. Reported perpetrators were varied and included colleagues (84%), patients (44%), staff (41%), administrators (18%), and trainees (16%). Prevalence of bullying (55.0 vs 47.7%, p=0.33), gender discrimination (59.1 vs 52.3%, p=0.33) and microaggressions (83.3 vs 83.0%, p=1.00) were similar irrespective of departmental leadership gender., Conclusions: Women gynecologic oncologists report a high prevalence of workplace bullying, gender discrimination and microaggressions regardless of the gender of their immediate leadership. Proactive and deliberate structural interventions to improve the work environment for surgeons who are women are urgently needed., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Hong, Rubinsak, Benoit, Teoh, Chandavarkar, Brockmeyer, Stevens, Ioffe and Temkin.)

This paper aims to develop a position tracking algorithm by which a rat in a radial arm maze can be accurately located in real time. An infrared (IR) night-vision camera was hung above the maze to capture IR images of the rat. The IR images were binarized and then duplicated for subsequent intersection and opening operations. Due to simple operations and a high robustness against the noise spots formed by the droppings of the rat, it took just minutes to process more than 9000 frames, and an accuracy above 99% was reached as well. The maze was intruded by an experimenter to further test the robustness, and the accuracy slightly fell to 98%. For comparison purposes, the same experiments were carried out using a pre-trained YOLO v2 model. The YOLO counterpart gave an accuracy beyond 97% in the absence and in the presence of the intruder. In other words, this work slightly outperformed the YOLO counterpart in terms of the accuracy in both cases, which indicates the robustness of this work. However, it took the YOLO counterpart an hour or so to locate a rat contained in the frames, which highlights the contribution of this work., (© 2021. The Author(s).)

Background: Adenocarcinoma in situ (AIS) of the cervix, is increasing in incidence, particularly in women of reproductive age. Fertility preservation is often desired. In a predominantly Hispanic population, we sought to determine the incidence of occult cervical cancer co-existing with AIS, and evaluate how conization margin status correlates with residual disease upon hysterectomy. Methods: A retrospective study utilizing a comprehensive cancer center database was conducted. Data from patients with histologically proven AIS of the cervix were abstracted. Results: Of 47 patients that met the criteria, 23 (49%) were Hispanic, 21 (45%) were White, two (4%) were Asian, and one (2%) was Black. The median age was 37. Forty-two patients underwent cervical conizations; 13/42 (48%) had positive margins upon conization; 28/42 (67%) underwent hysterectomies. Furthermore, 6/13 (46%) patients with positive conization margins had residual disease in hysterectomy specimens, with 2/13 (15%) found to have invasive cancer. In contrast, 0/14 (0%) of patients with negative margins had residual disease ( p = 0.036, Chi-squared 4.41, df = 1). In total, 2/27 (7%) patients who underwent hysterectomies had invasive cancer (7%). Conclusions: Positive margins upon cervical conization for AIS of the cervix were correlated with a relatively high rate of residual AIS and occult invasive cancer. Negative conization margins were correlated with no residual disease. Those patients may be candidates for fertility-sparing treatment.

Background: Patients with placenta accreta spectrum (PAS) disorders often suffer massive hemorrhage during cesarean hysterectomies (CHyst). A novel strategy to decrease blood loss and minimize perioperative morbidity associated with PAS is utilization of ER-REBOA Catheter intraoperatively. In this study, we explore the use of ER-REBOA Catheter during CHyst with the goal of minimizing perioperative morbidity and packed red blood cell (PRBC) transfusions., Methods: We conducted a retrospective case-control study at a regional referral center of consecutive patients with PAS undergoing CHyst. The primary outcomes were PRBC transfusions of ≥4 units. Secondary outcomes included surgical intensive care unit admissions, postoperative length of stay (LOS), postoperative ileus, and vascular complication rate. We also explored utilization of manual palpation and omission of precesarean fluoroscopy for resuscitative endovascular balloon occlusion of the aorta (REBOA) placement verification in distal aortic zone 3., Results: 90 patients were included in the study. REBOA and non-REBOA cases were similar in clinicodemographic characteristics. 17.7% of REBOA cases received ≥4 units of PRBC compared with 49.3% of non-REBOA cases (p=0.03). Zero REBOA patients developed postoperative ileus, whereas 18 (25%) non-REBOA patients did (p=0.02). LOS was reduced in the REBOA group. Postplacement fluoroscopy was omitted in all REBOA cases. Two postoperative arterial thrombotic events (2 of 19, 11% of REBOA patients) were identified in the REBOA group, one requiring a thrombectomy (1 of 19, 5%)., Discussion: Decrease in blood transfusions of ≥4 units of PRBC is demonstrated when ER-REBOA Catheter is placed in distal aortic zone 3 during CHyst performed for severe PAS disorders. The incidence of postoperative ileus and LOS are reduced in the ER-REBOA Catheter group. Placement and utilization of ER-REBOA Catheter during CHyst may be feasible without fluoroscopy when manual placement verification is performed by an experienced operator. Protocol modifications focusing on reducing thrombotic rate are ongoing., Level of Evidence: IV., Competing Interests: Competing interests: YJMI – consulting for Tesaro in 2018, consulting for Immunogen in 2018, consulting for AstraZeneca in 2019, consulting for Seagen Inc 2021., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

High-grade serous ovarian carcinoma (HGSOC) is the deadliest of gynecological cancers due to its high recurrence rate and acquired chemoresistance. RAS/MEK/ERK pathway activation is linked to cell proliferation and therapeutic resistance, but the role of MEK1/2-ERK1/2 pathway in HGSOC is poorly investigated. We evaluated MEK1/2 pathway activity in clinical HGSOC samples and ovarian cancer cell lines using immunohistochemistry, immunoblotting, and RT-qPCR. HGSOC cell lines were used to assess immediate and lasting effects of MEK1/2 inhibition with trametinib in vitro. Trametinib effect on tumor growth in vivo was investigated using mouse xenografts. MEK1/2 pathway is hyperactivated in HGSOC and is further stimulated by cisplatin treatment. Trametinib treatment causes cell cycle arrest in G1/0-phase and reduces tumor growth rate in vivo but does not induce cell death or reduce fraction of CD133+ stem-like cells, while increasing expression of stemness-associated genes instead. Transient trametinib treatment causes long-term increase in a subpopulation of cells with high aldehyde dehydrogenase (ALDH)1 activity that can survive and grow in non-adherent conditions. We conclude that MEK1/2 inhibition may be a promising approach to suppress ovarian cancer growth as a maintenance therapy. Promotion of stem-like properties upon MEK1/2 inhibition suggests a possible mechanism of resistance, so a combination with CSC-targeting drugs should be considered.

Transmission electron microscopy (TEM) is an all-in-one tool to visualize the complex systems of any specimen that is 1 nm in size or smaller. The current chapter provides detailed guidelines for imaging morphological changes during programmed cell necrosis using TEM as a single-step methodology. In this protocol, a novel aldehyde dehydrogenase inhibitor is used to induce cell programmed necrosis in ovarian cancer cell lines (A2780 and SKOV3). This process is followed by gradient dehydration with ethanol, chemical fixation, sampled grid preparation, and staining with 0.75% uranyl formate. Following fixation and grid preparation, cells are imaged using TEM. The resulting images reveal morphological changes consistent with necrotic morphology, including swelling of cells and organelles, appearance of vacuoles, and plasma membrane rupture followed by leakage of cellular contents. The current approach allows a single-step methodology for characterization of cell-programmed necrosis in cells based on morphology.

Objectives: The primary target is to reveal whether the resuscitation with hypertonic saline (HTS) or hydroxyethyl starch (HES) would have different effects on the myeloid-derived suppressor cell (MDSC) count and monocytic MDSC (M-MDSC)/granulocytic/neutrophilic MDSC (G-MDSC) rate in the peripheral blood, spleen, and bone marrow nucleated cells (BMNC) in a controlled hemorrhagic shock mouse model under secondary Escherichia coli bacterial infection attack, comparing to resuscitation with normal saline (NS) in 72 hours., Method: After hemorrhagic shock with bacteremia, which is induced by Escherichia coli bacterial infection attack, comparing to resuscitation with normal saline (NS) in 72 hours. Method . After hemorrhagic shock with bacteremia, which is induced by Escherichia coli 35218 injection, the mice were distributed into control, NS, HTS, and HES groups. The peripheral blood nucleated cells (PBNC), spleen single-cell suspension, and bone marrow nucleated cells were collected. The flow cytometry was used to detect the MDSC, M-MDSC, and G-MDSC., Result: In PBNC, after resuscitation with NS, the MDSC was continuously higher, while the rate of M-MDSC/G-MDSC were continuously lower ( P < 0.05). In HTS, the MDSC varied, higher at 24 and 72 hours ( P < 0.05). In HTS, the MDSC varied, higher at 24 and 72 hours ( P < 0.05). In HTS, the MDSC varied, higher at 24 and 72 hours ( P < 0.05). In HTS, the MDSC varied, higher at 24 and 72 hours ( P < 0.05). In HTS, the MDSC varied, higher at 24 and 72 hours ( P < 0.05), the M-MDSC/G-MDSC were continuously lower ( P < 0.05). In the spleen, resuscitation with HTS, the M-MDSC/G-MDSC were continuously lower ( P < 0.05). In BMNC, after resuscitation with HES, the M-MDSC/G-MDSC were lower at 24 and 72 hours ( P < 0.05)., Conclusion: In mouse hemorrhagic shock model with bacterial infection, the resuscitation with NS, HTS, or HES induced difference changes in MDSC and M-MDSC/G-MDSC, which were time-dependent and organ-specific. Resuscitation with crystalloid, like NS or HTS, showed longer effects on the MDSC and M-MDSC/G-MDSC in peripheral blood; while HTS has a longer effect on M-MDSC/G-MDSC in the spleen, HES has a stronger impact on the differentiation regulation of MDSC to G-MDSC in the bone marrow., Competing Interests: The authors have no conflict of interests to declare., (Copyright © 2020 Jiu-Kun Jiang et al.)

Prolonged occlusion of multiple microvessels causes microvascular injury. G protein-coupled receptor 124 (GPR124) has been reported to be required for maintaining central nervous system (CNS) angiogenesis and blood-brain barrier integrity. However, the molecular mechanisms by which GPR124 regulates pericytes during ischemia have remained elusive. Methods : A microsphere embolism-induced ischemia model was used to evaluate the expression of GPR124 following microsphere embolism. Immunocytochemistry and stochastic optical reconstruction microscopy imaging were used to assess the expression and distribution of GPR124 in human brain vascular pericytes (HBVPs) and after the treatment with 3-morpholino-sydnonimine (SIN-1) or oxygen-glucose deprivation (OGD). The effect of GPR124 knockdown or overexpression on HBVP migration was analyzed in vitro using wound healing assays and a microfluidic device. GPR124 loss-of-function studies were performed in HBVPs and HEK293 cells using CRISPR-Cas9-mediated gene deletion. Time-lapse imaging was used to assess dynamic changes in the formation of filopodia in an individual cell. Finally, to explore the functional domains required for GPR124 activity, deletion mutants were constructed for each of the N-terminal domains. Results : GPR124 expression was increased in pericytes following microsphere embolism. Morphological analysis showed localization of GPR124 to focal adhesions where GPR124 bound directly to the actin binding protein vinculin and upregulated Cdc42. SIN-1 or OGD treatment redistributed GPR124 to the leading edges of HBVPs where GPR124 signaling was required for pericyte filopodia formation and directional migration. Partial deletion of GPR124 domains decreased SIN-1-induced filopodia formation and cell migration. Conclusion : Taken together, our results provide the first evidence for a role of GPR124 in pericyte migration under ischemic conditions and suggest that GPR124 was essential for Cdc42 activation and filopodia formation., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Histone methylation is a modification which occurs in the N-terminal peptide chains of the histone nucleosome. The 4th, 9th, 27th, 36th and 79th lysines in N-terminal peptide chain of histone H3 are hot spots for this modification, including mono-, di-, and tri-methylation. H3K27me3 is the tri-methylation modification on histone H3 lysine 27, which mainly functions as a transcriptional repressor regulating skeletal muscle development. Studies have shown that H3K27me3 can finely regulate skeletal muscle proliferation, including the level and duration of skeletal muscle development by specifically binding to myogenic regulatory factors (e.g., MyoD, MyoG, etc.), cell cycling regulators, and epigenetic regulators including lncRNA and miRNA. In this review, we introduce the types and mechanisms of histone methylation and de-methylation of H3K27. We also summarize how H3K27me3 functions in the proliferation and differentiation of skeletal muscle cell. This review will contribute to the comprehension of the function of H3K27me3 in regulating skeletal muscle development and provide reference for further improving our understanding of mammalian muscle.

Aims: Autism spectrum disorder (ASD) is a wide range of neurodevelopmental disorders involving deficits in social interaction and communication. Unfortunately, autism remains a scientific and clinical challenge owing to the lack of understanding the cellular and molecular mechanisms underlying it. This study aimed to investigate the pathophysiological mechanism underlying leukocyte-endothelial adhesion in autism-related neurovascular inflammation., Methods: Male BTBR T+tf/J mice were used as an autism model. The dynamic pattern of leukocyte-endothelial adhesion in mouse cerebral vessels was detected by two-photon laser scanning microscopy (TPLSM). Using FACS, RT-PCR, and Western blotting, we explored the expression of cell adhesion molecules, the mRNA expression of endothelial chemokine, the protein levels of cathepsin B, and inflammatory mediators., Results: We found a significant increase in leukocyte-endothelial adhesion in BTBR mice, accompanied by elevated expression of the adhesion molecule neutrophils CD11b and endothelial ICAM-1. Our data further indicate that elevated neutrophil cathepsin B levels contribute to elevated endothelial chemokine CXCL7 levels in BTBR mice. The pharmacological inhibition of cathepsin B reverses the enhanced leukocyte-endothelial adhesion in the cerebral vessels of autistic mice., Conclusion: Our results revealed the prominent role of cathepsin B in modulating leukocyte-endothelial adhesion during autism-related neurovascular inflammation and identified a promising novel approach for autism treatment., (© 2018 John Wiley & Sons Ltd.)

Primary immune thrombocytopenia (ITP) is a bleeding disorder commonly encountered in clinical practice. The International Working Group (IWG) on ITP has published several landmark papers on terminology, definitions, outcome criteria, bleeding assessment, diagnosis, and management of ITP. The Chinese consensus reports for diagnosis and management of adult ITP have been updated to the 4th edition. Based on current consensus positions and new emerging clinical evidence, the thrombosis and hemostasis group of the Chinese Society of Hematology issued Chinese guidelines for management of adult ITP, which aim to provide evidence-based recommendations for clinical decision making.

Background/aims: Endothelial cell dysfunction is the principal pathological process underlying atherosclerotic cardiovascular disease. G protein-coupled receptor 124 (GPR124), an orphan receptor in the adhesion GPCR subfamily, promotes angiogenesis in the brain. In the present study, we explored the role of endothelial GPR124 in the development and progression of atherosclerosis in adult mice., Methods: Using tetracycline-inducible transgenic systems, we generated mice expressing GPR124 specifically under control of the Tie-2 promoter. The animal model of atherosclerosis was constructed by intravenously injecting AAV-PCSK9DY into tetracycline-regulated mice and feeding the mice a high-fat diet for 16 consecutive weeks. Biochemical analysis and immunohistochemistry methods were used to address the role and mechanism of GPR124 in the pathological process of atherosclerosis., Results: Higher TC (total cholesterol) and LDL-C (low density lipoprotein cholesterol) levels in serum and greater lipid deposition in the aortic sinus were found in atherosclerotic mice with GPR124 overexpression, coincident with the elevated proliferation of smooth muscle cells. We observed an elevation of ONOO- in the aortic sinus in this model by using immunofluorescence, and the experiments showed that the specific overexpression of GPR124 in the endothelium induced the up-regulation of CD68, NLRP3 and caspase-1 levels in the aortic sinus., Conclusion: The above results indicate that manipulating GPR124 in the endothelium may contribute to delayed pathological progression of atherosclerosis., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

Objective: Melanoma originating from gynecologic sites (MOGS), including the vulva, vagina, and cervix, is a rare and aggressive form of melanoma with poor long-term clinical outcome. The clinicopathologic features of vulvar and non-vulvar tumors remain relatively understudied, and in contrast to cutaneous melanomas at non-sun-exposed sites, MOGS typically do not harbor BRAF mutations. Thus, we sought to analyze the clinicopathologic and molecular features of MOGS., Methods: A large retrospective cohort of patients with MOGS (n=59) at a single large academic institution over a 28-year period was identified. Associations among clinicopathologic characteristics were assessed via standard statistical approaches, and clinical outcome was examined using Cox regression analysis. Sanger sequencing was utilized to identify mutations in hotspot regions of BRAF, KIT, NRAS, and CTNNB1., Results: Tumors involving the vagina and/or cervix (non-vulvar) are significantly associated with high-risk clinicopathologic features, including increased tumor thickness, ulceration, positive resection margins, lymph node metastasis, and poor long-term clinical outcome (with increased risk of death due to disease). The aggressive clinical behavior of non-vulvar tumors is independent of advanced clinical stage and lymph node metastasis in multivariate analysis. Targeted molecular analysis confirms an overall low rate of oncogenic mutations in our MOGS cohort, although KIT mutations (particularly in exon 11) are relatively enriched., Conclusions: Overall, our results show that non-vulvar MOGS are aggressive tumors with poor long-term clinical outcome and indicate that few targeted therapeutic options are currently available to patients with MOGS., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective: To explore the stability of resistant phenotypes and changes of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) gene system on four Shigella strains in the absence of antibiotics. Methods: Four clinical isolated Shigella strains that resistant to different antibiotics were consecutive passaged for 90 times without antibiotics. Agar dilution method was used to determine the minimum inhibitory concentration of Shigella strains. After sequence analysis with PCR, CRISPR Finder and Clustal X 2.1 were applied to identify the changes of CRISPR loci in the Shigella strains. Results: After the consecutive transfer of 90 generations, sensitivity to certain antibiotics of four Shigella strains with different drug resistant spectrums increased. Mel-sf1998024/zz resistance to ampicillin, cephalexin, cefotaxime, chloramphenicol decreased, mel-s2014026/sx resistance to norfloxacin, trimethoprim decreased, mel-sf2004004/sx drug resistance to ampicillin, cefuroxime, cefotaxime, chloramphenicol, trimethoprim decreased and mel-sf2013004/bj resistance to chloramphenicol decreased. The spacer of which matched gene codes Cas and its upstream repeat in 3'end of CRISPR3 got lost in mel-sf1998024/zz and mel-sf2013004/bj. Conclusions: Shigella strains could reduce or lose their resistance to some antibiotics after consecutive transfers, without the interference of antibiotics. CRISPR3 locus had dynamic spacers in Shigella strains while CRISPR3 locus and cas genes might have been co-evolved.

Objective: To investigate the distribution and differentiation of myeloid-derived suppressor cells (MDSCs) in hemorrhagic shock mice, which are resuscitated with normal saline (NS), hypertonic saline (HTS), and hydroxyethyl starch (HES)., Methods: BALB/c mice were randomly divided into control, NS, HTS, and HES resuscitation groups. Three subgroups (n=8) in each resuscitation group were marked as 2, 24, and 72 h. Flow cytometry was used to detect the MDSCs, monocytic MDSCs (M-MDSCs), and granulocytic/neutrophilic MDSCs (G-MDSCs) in peripheral blood nucleated cells (PBNCs), spleen single-cell suspension, and bone marrow nucleated cells (BMNCs)., Results: The MDSCs in BMNCs among three resuscitation groups were lower 2 h after shock, in PBNCs of the HTS group were higher, and in spleen of the NS group were lower (all P<0.05 vs. control). The M-MDSC/G-MDSC ratios in PBNCs of the HTS and HES groups were lower (both P<0.05 vs. control). At 24 h, the MDSCs in PBNCs of the NS and HTS groups were higher, while the spleen MDSCs in the HTS group were higher (all P<0.05 vs. control). The M-MDSC/G-MDSC ratios were all less in PBNCs, spleen, and BMNCs of the NS and HTS groups, and were lower in BMNCs of the HES group (all P<0.05 vs. control). At 72 h, the elevated MDSCs in PBNCs were presented in the HTS and HES groups, and in spleen the augment turned up in three resuscitation groups (all P<0.05 vs. control). The inclined ratios to M-MDSC were exhibited in spleen of the NS and HTS groups, and in PBNCs of the NS group; the inclination to G-MDSC in BMNCs was shown in the HES group (all P<0.05 vs. control)., Conclusions: HTS induces the earlier elevation of MDSCs in peripheral blood and spleen, and influences its distribution and differentiation, while HES has a less effect on the distribution but a stronger impact on the differentiation of MDSCs, especially in bone marrow., Competing Interests: Compliance with ethics guidelines: Jiu-kun JIANG, Wen FANG, Liang-jie HONG, and Yuan-qiang LU declare that they have no conflict of interest. All institutional and national guidelines for the care and use of laboratory animals were followed.

Objective: We conducted this study to analyze the effects of nucleoside analogues in the treatment of hepatitis B cirrhosis and its effect on Th17 cells., Patients and Methods: 120 patients were randomly divided into lamivudine combined with adefovir dipivoxil group (combined group) and entecavir group. There were 59 cases in the combined group and 59 cases in entecavir group. The combined group was administered lamivudine 100 mg/d + adefovir dipivoxil 10 mg/d and entecavir group was administered entecavir 0.5 mg/d. The treatment was continued until there was virus negativity and it maintains for at least 3 months., Results: The treatment effects were compared. We compared the average rate of viral clearance time and virus clearance of two groups of patients; the difference was not statistically significant (p>0.05). The relapse rate after a negative test of entecavir group was lower than that of the combined group (p<0.05). Before and after treatment, the levels of TBIL, ALT and ALB in the two groups were compared; the differences were not statistically significant (p>0.05). The Th17 cell proportion and the level of IL-17 after treatment of the entecavir group were lower than those before treatment. The combined group exhibited no change, and the entecavir group was lower than combined group; the differences were statistically significant (p<0.05)., Conclusions: Therefore, the effects of the combination of lamivudine and adefovir dipivoxil is the same as single entecavir treatment of hepatitis B cirrhosis suppression of viral replication. It does not increase liver injury and the antiviral effects of entecavir may be related to inhibition of the expression of Th17 cells and effector molecules IL-17.

Objective: A new method related to molecular biomarker with CRISPR/Cas (clustered regularly interspaced short palindromic repeats-cas) in Escherichia (E.) coli was developed and used for surveillance programs., Methods: CRISPR/Cas sequence that containing 135 strains with complete sequence and 203 strains with whole genome shotgun sequence of E. coli in GenBank by BLAST and 361 strains of E. coli (including 38 strains of E. coli O157∶H7) in laboratory were identified by PCR and analyzed with the CRISPR Finder. Spacers were compared with DANMAN and the phylogenetic trees of cas gene were constructed under Clustal Ⅹ and Mega 5.1., Results: With new perspective, a descriptive method was developed targeting on the position of CRISPR/cas in E. coli. The CRISPR1 was detected in 77.04%, 100.00% and 75.62% and the CRISPR2 was detected in 74.81%, 100.00% and 92.24% and the CRISPR3 and CRISPR4 were detected in 11.85%, 0 and 1.39% for 135 strains with complete sequence, 203 strains with whole genome shotgun sequence and 361 strains in the laboratory, respectively. One strain downloaded in GenBank with whole genome sequencing and 2 strains in the our laboratory were identified that containing four CRISPR locus. The other E. coli strain was with insertion sequence in downstream of the non-cas CRISPR1. The unique CRISPR was found in 8 strains of O55∶H7, in 180 strains of O157∶H7, in 8 strains of O157∶HNM, in 40 strains of O104∶H4, in 4 strains of O145∶H28, in all the 699 E. coli strains. The phylogenetic tree could be divided into two groups-cas with type I-E or type I-F., Conclusions: CRISPR/Cas might be used as a valuable molecular biomarker in epidemiological surveillance studies to identify the high virulent strains or new strains of E. coli. Phage night be related to the missing or obtaining of spacers.

Valproate exposure is associated with increased risks of autism spectrum disorder. To date, the mechanistic details of disturbance of melatonin receptor subtype 1 (MTNR1A) internalization upon valproate exposure remain elusive. By expressing epitope-tagged receptors (MTNR1A-EGFP) in HEK-293 and Neuro-2a cells, we recorded the dynamic changes of MTNR1A intracellular trafficking after melatonin treatment. Using time-lapse confocal microscopy, we showed in living cells that valproic acid interfered with the internalization kinetics of MTNR1A in the presence of melatonin. This attenuating effect was associated with a decrease in the phosphorylation of PKA (Thr197) and ERK (Thr202/Tyr204). VPA treatment did not alter the whole-cell currents of cells with or without melatonin. Furthermore, fluorescence resonance energy transfer imaging data demonstrated that valproic acid reduced the melatonin-initiated association between YFP-labeled β-arrestin 2 and CFP-labeled MTNR1A. Together, we suggest that valproic acid influences MTNR1A intracellular trafficking and signaling in a β-arrestin 2-dependent manner.

Accumulating evidence suggests that formation of peroxynitrite (ONOO(-)) in the cerebral vasculature contributes to the progression of ischemic damage, while the underlying molecular mechanisms remain elusive. To fully understand ONOO(-) biology, efficient tools that can realize the real-time tracing of endogenous ONOO(-) fluxes are indispensable. While a few ONOO(-) fluorescent probes have been reported, direct visualization of ONOO(-) fluxes in the cerebral vasculature of live mice remains a challenge. Herein, we present a fluorescent switch-on probe (NP3) for ONOO(-) imaging. NP3 exhibits good specificity, fast response, and high sensitivity toward ONOO(-) both in vitro and in vivo. Moreover, NP3 is two-photon excitable and readily blood-brain barrier penetrable. These desired photophysical and pharmacokinetic properties endow NP3 with the capability to monitor brain vascular ONOO(-) generation after injury with excellent temporal and spatial resolution. As a proof of concept, NP3 has enabled the direct visualization of neurovascular ONOO(-) formation in ischemia progression in live mouse brain by use of two-photon laser scanning microscopy. Due to these favorable properties, NP3 holds great promise for visualizing endogenous peroxynitrite fluxes in a variety of pathophysiological progressions in vitro and in vivo.

Optical design in enhancing optical absorption of group-III-nitride- and multiple quantum well-based GaN/InxGa 1- xN/cSi dual-junction tandem solar cells with triangular diffraction grating is simulated and optimized by using combined two-dimensional rigorous coupled wave analysis and transfer matrix methods. This paper thoroughly examines these phenomena of optical absorption affected by antireflection coatings, multiple thin-film layers and diffraction gratings with the integrated perspectives of semiconductor physics and electromagnetic theory for the first time. An improvement of 58% in absorption compared to the prototype SC is obtained which means more than 80% of incoming light (hυ > Eg Si ) can be harvested in this thin-film (< 4 μm in total) design.

Septic encephalopathy (SE) is a critical factor determining sepsis mortality. Vascular inflammation is known to be involved in SE, but the molecular events that lead to the development of encephalopathy remain unclear. Using time-lapse in vivo two-photon laser scanning microscopy, we provide the first direct evidence that cecal ligation and puncture in septic mice induces microglial trafficking to sites adjacent to leukocyte adhesion on inflamed cerebral microvessels. Our data further demonstrate that septic injury increased the chemokine CXCL1 level in brain endothelial cells by activating endothelial P2RX7 and eventually enhanced the binding of Mac-1 (CD11b/CD18)-expressing leukocytes to endothelial ICAM-1. In turn, leukocyte adhesion upregulated endothelial CX3CL1, thereby triggering microglia trafficking to the injured site. The sepsis-induced increase in endothelial CX3CL1 was abolished in CD18 hypomorphic mutant mice. Inhibition of the P2RX7 pathway not only decreased endothelial ICAM-1 expression and leukocyte adhesion but also prevented microglia overactivation, reduced brain injury, and consequently doubled the early survival of septic mice. These results demonstrate the role of the P2RX7 pathway in linking neurovascular inflammation to brain damage in vivo and provide a rationale for targeting endothelial P2RX7 for neurovascular protection during SE.

Background: A viable tubal pregnancy is an extremely rare occurrence with an increased risk of fetal as well as maternal morbidity and mortality. We report a third trimester tubal pregnancy occurring after an interval tubal ligation., Case: A 43-year-old woman, gravida 2 para 1, presented at 29 weeks' gestation with an asymptomatic extrauterine pregnancy and was managed expectantly in the antepartum unit. At 33 weeks the fetus was delivered for worsening umbilical artery velocimetry. Despite aggressive resuscitative efforts, the neonate did not survive., Conclusion: In managing an advanced extrauterine pregnancy, imaging with MRI may help diagnose and confirm suspicion raised by ultrasonography and may aid in presurgical planning and management. This case illustrates the diagnostic challenge and high neonatal mortality of an advanced tubal pregnancy.

The present study was designed to investigate the role of autophagy-lysosome signaling in the brain after application of nanoparticles. Here, lipid nanoparticles (LNs) induced elevations of Atg5, P62, LC3 and cathepsin B in mice brain. The transmission electron microscopy revealed a dramatic elevation of lysosome vacuoles colocalized with LNs cluster inside the neurons in mice brain. Immunoblot data revealed abnormal expression of cathepsin B in brain cortex following LNs injection, whereas its expression was further elevated in Atg5(+/-) mice. The importance of Atg5 in the LNs-induced autophagy-lysosome cascade was further supported by our finding that neurovascular response was exaggerated in Atg5(+/-) mice. In addition, the siRNA knockdown of Atg5 significantly blunted the increasing of LC3 and P62 in LNs-treated Neuro-2a cells. Taken together, we propose that LNs induce autophagy-lysosome signaling and neurovascular response at least partially via an Atg5-dependent pathway., From the Clinical Editor: These authors investigated autophagy-lysosome signaling in the mouse brain after application of lipid nanoparticles and report that these nanoparticles induce autophagy-lysosome signaling and neurovascular response at least partially via an Atg5-dependent pathway., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Aims: Diabetes mellitus increases the risk of stroke, but the mechanisms are unclear. The present study tested the hypothesis that diabetes mellitus disturbs the brain microcirculation and increases the susceptibility to cerebral damage in a middle cerebral artery occlusion (MCAO) model of ischemia., Methods: Diabetes was induced by streptozocin in mice expressing green fluorescent protein in endothelial cells (Tie2-GFP mice). Four weeks later, they were subjected to transient (20 min) MCAO. In vivo blood flow was measured by two-photon laser-scanning microscopy (TPLSM) in cerebral arteries, veins, and capillaries., Results: There was a significant decrease in red blood cell (RBC) velocity in capillaries in diabetic mice as assessed by TPLSM, yet the regional cerebral blood flow, as assessed by laser Doppler flowmetry, was maintained. Brain capillary flow developed turbulence after MCAO only in diabetic mice. These mice sustained increased neurological deficits after MCAO which were accompanied by an exaggerated degradation of tight junction proteins and blunted CaMKII phosphorylation in cerebral tissues indicating disruption of the blood-brain barrier and disturbed cognitive potential., Conclusion: Diabetic mice are more susceptible to disturbances of cerebral capillary blood flow which may predispose them to neurovascular defects following ischemia., (© 2014 John Wiley & Sons Ltd.)

Aims: Although there is accumulating evidence that increased formation of reactive nitrogen species in cerebral vasculature contributes to the progression of ischemic damage, but the underlying molecular mechanisms remain elusive. Peroxiredoxin 1 (Prx1) can initiate the antioxidant response by scavenging free radicals. Therefore, we tested the hypothesis that Prx1 regulates the susceptibility to nitrosative stress damage during cerebral ischemia in vitro and in vivo., Results: Proteomic analysis in endothelial cells revealed that Prx1 was upregulated after stress-related oxygen-glucose deprivation (OGD). Although peroxynitrite upregulated Prx1 rapidly, this was followed by its polyubiquitination within 6 h after OGD mediated by the E3 ubiquitin ligase E6-associated protein (E6AP). OGD colocalized E6AP with nitrotyrosine in endothelial cells. To assess translational relevance in vivo, mice were studied after middle cerebral artery occlusion (MCAO). This was accompanied by Prx1 ubiquitination and degradation by the activation of E6AP. Furthermore, brain delivery of a lentiviral vector encoding Prx1 in mice inhibited blood-brain barrier leakage and neuronal damage significantly following MCAO., Innovation and Conclusions: Nitrosative stress during ischemic insult activates E6AP E3 ubiquitin ligase that ubiquitinates Prx1 and subsequently worsens cerebral damage. Thus, targeting the Prx1 antioxidant defense pathway may represent a novel treatment strategy for neurovascular protection in stroke.

The translation of experimental stroke research from the laboratory to successful clinical practice remains a formidable challenge. We previously reported that PEGylated-lipid nanoparticles (PLNs) effectively transport across the blood-brain barrier along with less inflammatory responses. In the present study, PLNs conjugated to Fas ligand antibody that selectively present on brain ischaemic region were used for therapeutic targeting. Fluorescent analysis of the mice brain show that encapsulated 3-n-Butylphthalide (dl-NBP) in PLNs conjugated with Fas ligand antibody effectively delivered to ipsilateral region of ischaemic brain. Furthermore, the confocal immunohistochemical study demonstrated that brain-targeted nanocontainers specifically accumulated on OX42 positive microglia cells in ischaemic region of mice model. Finally, dl-NBP encapsulated nano-drug delivery system is resulted in significant improvements in brain injury and in neurological deficit after ischaemia, with the significantly reduced dosages versus regular dl-NBP. Overall, these data suggests that PLNs conjugated to an antibody specific to the Fas ligand constituted an ideal brain targeting drug delivery system for brain ischaemia., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Lower global cognitive function scores are a common symptom of autism spectrum disorders (ASDs). This study investigates the effects of melatonin on hippocampal serine/threonine kinase signaling in an experimental ASD model. We found that chronic melatonin (1.0 or 5.0 mg/kg/day, 28 days) treatment significantly rescued valproic acid (VPA, 600 mg/kg)-induced decreases in CaMKII (Thr286), NMDAR1 (Ser896), and PKA (Thr197) phosphorylation in the hippocampus without affecting total protein levels. Compared with control rats, the immunostaining of pyramidal neurons in the hippocampus revealed a decrease in immunolabeling intensity for phospho-CaMKII (Thr286) in the hippocampus of VPA-treated rats, which was ameliorated by chronic melatonin treatment. Consistent with the elevation of CaMKII/PKA/PKC phosphorylation observed in melatonin-treated rat, long-term potentiation (LTP) was enhanced after chronic melatonin (5.0 mg/kg) treatment, as reflected by extracellular field potential slopes that increased from 56 to 60 min (133.4 ± 3.9% of the baseline, P < 0.01 versus VPA-treated rats) following high-frequency stimulation (HFS) in hippocampal slices. Accordingly, melatonin treatment also significantly improved social behavioral deficits at postnatal day 50 in VPA-treated rats. Taken together, the increased phosphorylation of CaMKII/PKA/PKC signaling might contribute to the beneficial effects of melatonin on autism symptoms., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Nanocarrier-based drug delivery systems have attracted wide interest for the treatment of brain disease. However, neurotoxicity of nanoparticle has limited their therapeutic application. Here we demonstrated that lipid nanoparticles (LNs) accumulated in the brain parenchyma within 3 h of intravenous injection to mice and persisted for more than 24 weeks, coinciding with a dramatic activation of brain microglia. Morphological characteristic of microglial activation also observed in LNs-treated Cx3cr1GFP/+ mice. In vivo study with two-photon confocal microscopy revealed abnormal Ca²⁺ waves in microglia following LNs injection. The correlated activation of caspase-1, IL-1β and neurovascular damage following LNs injection was attenuated in P2X₇-/- mice. PEGylation of LNs reduced correlated nanoparticles aggregation. Moreover, PEGylation of LNs ameliorated the P2X₇/caspase-1/IL-1β signalling-dependent microglia activation and neurovascular damage. In conclusion, PEGylation of LNs is a promising biomaterial for brain-targeted therapy that inhibits P2X7₇-dependent neuroinflammatory response., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Purpose: Placentas from spontaneous abortions and preterm deliveries have a higher prevalence of Human papillomavirus (HPV) compared to placentas from elective abortions and term births. The objective was to determine the effects of HPV-16 on the adhesion and implantation properties of early embryo trophoblasts., Methods: Two-cell mouse embryos were cultured (medium G2, 5 % CO2, 37 °C) for 72-96 h and exposed to either HPV-16 rich SiHa cell lysates which were refrigerated after mechanical lysis, thawed lysates which had been frozen for freeze/thaw lysis method, or control medium, incubated (4-5 days) and evaluated by microscopy (N = 96 embryos, 3 repeated experiments). Trophoblasts were stained and images were digitized. Adhesion and dimension data were analyzed by Chi-square and t test, respectively., Results: HPV-16 exposed embryos exhibited less adhesion through reduced implantation compared with the control (combined lysates 53.8 vs. 85.7 %, P < 0.05). Refrigerated and thawed lysate groups had similar reduced implantations (58.3 vs. 50.0 %). Of the embryos with implantation, 100 % in the refrigerated lysates were noted to have loose or abnormal adhesion. This was measured when embryos were noted to be lost after washes with HTF. There was no difference in trophoblast viability among the groups. Total trophoblast area was greater in the HPV-16 exposed frozen lysate group (1,881.8 ± 605.3 vs. control 848.8 ± 298.0 square units, mean ± SEM)., Conclusions: HPV-16 inhibited trophoblasts adhesion needed for normal implantation, but not embryo development. Total trophoblast spread was increased after HPV-16 exposure suggesting that HPV-16 altered trophoblast migration. These results suggest that HPV-16 may induce abnormal placental growth resulting in pregnancy wastage.

Thirteen kinds of common drug precursor chemicals were detected on the spot by transportable Raman spectrometer independently researched and developed. Changing analysis methods, such as the focal position of the instrument, analysis time, external light source, sample containers and the concentration of the analyte, the high quality Raman spectrums were obtained. The study shows that analysis method can be optimized to improve the detection efficiency, LOD (limit of detection) of 1-phenyl-2-acetone, toluene and chloroform was recorded for lower concentrations and the component contents of the mixture have a significant impact on the test results. Selecting the appropriate instrument conditions can be more effective and fast screening of precursor chemicals on site to provide help for those who are engaged in the anti-drug work.

Three new triterpenoid saponins, tomentoside A (1), B (2) and C (3), along with four known saponins (4-7) were isolated from the root of Anemone tomentosa. The structures of the new compounds were elucidated as 3-O-β-D-ribopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-[β-D-glucopyranosyl-(1→4)]-α-L-arabinopyranosyl hederagenin 28-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (1), 3-O-β-D-ribopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-[β-D-glucopyranosyl-(1→4)]-β-D-xylopyranosyl hederagenin 28-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (2) and 3-O-β-D-galactopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranosyl oleanolic acid 28-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranosyl-(1→6)-β-D-glucopyranoside (3) on the basis of chemical and spectral evidence. In the oligosaccharide chains of compound 3, the characteristic D-galactose residue is a rare structural feature and secondly encountered among triterpenoid saponins from Anemone.

A solid-phase extraction (SPE)-high performance liquid chromatography (HPLC) method has been developed for simultaneous determination of 10 sulfonamide antibiotics in water. The analytes were first enriched and purified through a PEP solid-phase extraction column, and eluted with acetonitrile-dichloromethane solution (2: 1, V/V), then detected by a HPLC with a UV detector. The detection wavelength was 268 nm and the column temperature was 33 degrees C, using gradient elution process with acetonitrile - 0.4% acetic acid/water (V/V) as the mobile phase to achieve baseline separations of these 10 analytes. The linearity range was 10 - 2 000 microg x L(-1). The recovery ranges of standard addition for deionized water and real water samples were 73.4% - 95.6% and 70.2% - 92.5%, respectively (except for sulfonamide, were 8.5% and 8.0%). The limit of detection was 1.42-7.25 ng x L(-1). Application of this method for parts of Huangpu River in Shanghai, surface water and groundwater in Chongming Island showed that sulfonamide antibiotics were detected in different frequencies in different aqueous environments, with the concentration range of 13.3 - 241.5 ng x L(-1), proving this method is easy, rapid, sensitive and efficient to meet the needs of actual work.

The Escherichia coli Lon protease degrades the E. coli DNA-binding protein HUbeta, but not the related protein HUalpha. Here we show that the Lon protease binds to both HUbeta and HUalpha, but selectively degrades only HUbeta in the presence of ATP. Mass spectrometry of HUbeta peptide fragments revealed that region K18-G22 is the preferred cleavage site, followed in preference by L36-K37. The preferred cleavage site was further refined to A20-A21 by constructing and testing mutant proteins; Lon degraded HUbeta-A20Q and HUbeta-A20D more slowly than HUbeta. We used optical tweezers to measure the rupture force between HU proteins and Lon; HUalpha, HUbeta, and HUbeta-A20D can bind to Lon, and in the presence of ATP, the rupture force between each of these proteins and Lon became weaker. Our results support a mechanism of Lon protease cleavage of HU proteins in at least three stages: binding of Lon with the HU protein (HUbeta, HUalpha, or HUbeta-A20D); hydrolysis of ATP by Lon to provide energy to loosen the binding to the HU protein and to allow an induced-fit conformational change; and specific cleavage of only HUbeta., (Copyright 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Although the effect of mouse resistin on insulin-resistance has been well defined, but the biological function of human resistin is still unknown. This study was aimed to explore the possible physiological and pathological effects of human resistin, as well as the tissue distribution of human resistin and correlation of resistin gene expression with leukemia incidence. 152 leukemia patients without inflammatory complication and 100 healthy persons were selected as experimental and control groups respectively. The blood samples were collected, the total RNA was extracted, the expression distribution of resistin in different tissues was detected by semi-quantitative RT-PCR and then the statistical analysis was carried out. The results indicated that the expression of the human resistin gene was detected in normal fetus liver, adult bone marrow and umbilical cord blood and peripheral blood cells, while the resistin gene could not be amplified in fat, umbilical cord, placenta and adult liver. The resistin expression was detected in 21% leukemia patients and 27% healthy persons. The difference of the resistin gene expression between the two groups was not statistically significant (p>0.05). It is concluded that the higher expression of resistin exists in normal human fetus liver, adult bone marrow, umbilical cord blood and peripheral blood cells, which indicates that the distribution of human resistin correlates with normal hematopoiesis in certain extent, but its expression level and rate may not correlate with the incidence of leukemia.

The aim of this study was to evaluate the efficacy of recombinant adenovirus p53 agent (rAd-p53) injection combined with cisplatin (CDDP) for the treatment of malignant pleural or peritoneal effusion. After puncture drainage, patients in the treatment group (n = 27) received intracavitary administration of rAd-p53 (2 x 10(12) virus particles) once a week for 4 weeks. At 48 h after each rAd-p53 injection, patients were given intracavitary administration of cisplatin 60 mg/m(2). This administration procedure continued once a week for 4 weeks. The control group (n = 21) received the same intracavitary therapy as the treatment group but without rAd-p53 therapy. Efficacy was evaluated by clinical observations, computed tomography, tumour markers, Karnofsky score and short-term follow-up. The total effective rates for the treatment group (63.0%) were significantly higher than for the control group (42.9%), suggesting that the treatment group benefited over the control group. In conclusion, rAd-p53 therapy is a safe and effective treatment for advanced malignant pleural or peritoneal effusion.

The present study aimed for a clear visualization of faintly deposited colloidal iron in tissue sections for light microscopy. Paraffin blocks containing paraformaldehyde-fixed brain tissue from healthy adult mice were cut into sections 10-15 microm thick. After deparaffinization, the sections were stained with fine cationic iron colloid at a pH value of 1.0-1.5, and treated with a mixture of potassium ferrocyanide and hydrochloride for Prussian blue reaction. Some sections were further treated with Bodian's protein silver after the Prussian blue reaction. This sensitized development of Prussian blue reaction with Bodian's protein silver more clearly visualized the faintly deposited cationic colloidal irons than the demonstration by Prussian blue reaction alone, and allowed an enhanced visualization of the perineuronal nets of sulfated proteoglycans in the brain. Thus, such fine perineuronal sulfated proteoglycans as those in the CA3 field of the hippocampus, which are weakly stained with cationic iron colloid and usually overlooked by a demonstration with only a Prussian blue reaction, could be clearly visualized with striking contrast by the sensitized development with Bodian's protein silver after the Prussian blue reaction. Preliminary hyaluronidase digestion erased Bodian's protein silver development of perineuronal sulfated proteoglycans. Though some axonal fibers were also additionally stained with Bodian's protein silver itself, this sensitized development is useful to enhance such weak colloidal iron signals as are hardly detectable by only Prussian blue reaction.

Sections of the visual cortex of newborn (1-4 weeks after birth) and adult cats were stained with cationic iron colloid, aldehyde fuchsin or lectins (lectin Vicia villosa, soybean and Wisteria floribunda agglutinins). Many neurons in the adult cat visual cortex contained perineuronal sulfated proteoglycans detectable with cationic iron colloid and aldehyde fuchsin, or cell surface glycoproteins reactive to lectins. Double staining indicated that some of the lectin-labeled neurons were not stained with cationic iron colloid, and also that some of the cationic iron colloid-stained neurons were not labeled with lectins. The perineuronal sulfated proteoglycans and cell surface glycoproteins developed 3 weeks after birth. In the newborn cats 1-2 weeks after birth, no neurons were reactive to cationic iron colloid, aldehyde fuchsin or lectins. In the newborn cats 3-4 weeks after birth, it was clearly observed that the cytoplasm of the glial cells closely associated with the neurons containing the perineuronal sulfated proteoglycans showed an intense reaction to cationic iron colloid and aldehyde fuchsin, and that the Golgi complexes of the neurons with cell surface glycoproteins were intensely labeled with lectins. These findings suggest that the perineuronal sulfated proteoglycans are derived from the associated glial cells, and that the cell surface glycoproteins are produced by the associated nerve cells.

Sections of the retrosplenial cortex from adult and newborn mouse brains were observed with a light microscope. The retrosplenial cortex of the adult animals contained many neurons (10% of the total), including some dark neurons, with perineuronal sulfated proteoglycans detectable with cationic iron colloid and aldehyde fuchsin. The retrosplenial cortex of the adult animals also contained many neurons (10% of the total) with cell surface glycoproteins reactive to lectin Vicia villosa, soybean or Wisteria floribunda agglutinin. Double staining showed that the majority (75%) of the neurons labeled with lectins were stained with cationic iron colloid, and that some (25%) of them were not stained with this colloid. Double staining also showed that some (25%) of the neurons stained with cationic iron colloid were not labeled with lectins. These findings indicate that the perineuronal sulfated proteoglycans are, at least partly, independent from the cell surface glycoproteins. Observations of the sections from the newborn animals revealed that the perineuronal sulfated proteoglycans were produced by the associated satellite astrocytes 3-4 weeks after birth, and that the cell surface glycoproteins were produced by the associated nerve cells at earlier stages, or 2-3 weeks after birth. Dark neurons began to appear 3-4 weeks after birth. These dark neurons or their Golgi complexes were also reactive to lectins, suggesting the production of cell surface glycoproteins.

The present study was undertaken to investigate whether less pathogenic Candida species (C. tropicalis, C. stellatoidea, C. krusei and C. glabrata) express a fibronectin receptor (FNr) antigenically related to alpha 5 beta 1 integrin, which mediates their binding to fibronectin (FN). By flow cytometric analysis, a monoclonal antibody (MAb) directed against human alpha 5 integrin subunit (clone SAM-1) and two different antisera to FNr positively stained C. tropicalis, C. stellatoidea and C. glabrata, with the greatest expression observed for C. tropicalis. No or only marginal immunoreactivity was found on C. krusei. C. tropicalis, C. stellatoidea, C. glabrata, but not C. krusei yeasts specifically adhered to FN; higher levels of adhesion were found for C. tropicalis and C. stellatoidea with respect to C. glabrata. Less pathogenic Candida spp. bound to the Arg-Gly-Asp (RGD) containing 120-kDa fragment of FN and adhesion to intact FN was markedly inhibited by Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP), but not by Gly-Arg-Gly-Glu-Ser-Pro (GRGESP) peptides. In addition, anti-alpha 5 SAM-1 MAb and both anti-FNr antisera strongly blocked binding of less pathogenic Candida spp. to FN. Overall, these results indicate that less pathogenic Candida spp., including C. tropicalis, C. stellatoidea and C. glabrata, express a receptor antigenically related to alpha 5 beta 1 integrin which mediates their adhesion to FN.

The inhabitants with positive eggs of whipworm and ascaris in a village of Jiangdou County were examined and treated with different antihelmintic drugs twice a year. The uncured population were treated by drug administration once again. On an average the efficiency of drug treatment (cure rate x treatment rate) was 56.7% in whipworm and 64.5% in ascaris. The program had been carried out for five years. The positive rate of eggs of whipworm and ascaris in the population decreased from 64.7% and 52.1% to 2.7% and 4.1%, respectively, the mean increase in infection rates among population at 5 or 7 months after treatment was 2.68 (0.86-5.74)% and 6.94(1.72-17.27)%, respectively.