A population of descending tyraminergic/octopaminergic projection neurons of the insect deutocerebrum

In this study, we describe a cluster of tyraminergic/octopaminergic neurons in the lateral dorsal deutocerebrum of desert locusts (Schistocerca gregaria) with descending axons to the abdominal ganglia. In the locust, these neurons synthesize octopamine from tyramine stress-dependently. Electrophysiological recordings in locusts reveal that they respond to mechanosensory touch stimuli delivered to various parts of the body including the antennae. A similar cluster of tyraminergic/octopaminergic neurons was also identified in the American cockroach (Periplaneta americana) and the pink winged stick insect (Sipyloidea sipylus). It is suggested that these neurons release octopamine in the ventral nerve cord ganglia and, most likely, convey information on arousal and/or stressful stimuli to neuronal circuits thus contributing to the many actions of octopamine in the central nervous system.     

Combined use of cytology,p16 immunostaining and genotyping for triage of women positive for high-risk human papillomavirus at primary screening

Human papillomavirus (HPV) testing is very sensitive for primary cervical screening but has low specificity. Triage tests that improve specificity but maintain high sensitivity are needed. Women enrolled in the experimental arm of Phase 2 of the New Technologies for Cervical Cancer randomized controlled cervical screening trial were tested for high-risk HPV (hrHPV) and referred to colposcopy if positive. hrHPV-positive women also had HPV genotyping (by polymerase chain reaction with GP5+/GP6+ primers and reverse line blotting), immunostaining for p16 overexpression and cytology. We computed sensitivity, specificity and positive predictive value (PPV) for different combinations of tests and determined potential hierarchical ordering of triage tests. A number of 1,091 HPV-positive women had valid tests for cytology, p16 and genotyping. Ninety-two of them had cervical intraepithelial neoplasia grade 2+ (CIN2+) histology and 40 of them had CIN grade 3+ (CIN3+) histology. The PPV for CIN2+ was >10% in hrHPV-positive women with positive high-grade squamous intraepithelial lesion (61.3%), positive low-grade squamous intraepithelial lesion (LSIL+) (18.3%) and positive atypical squamous cells of undetermined significance (14.8%) cytology, p16 positive (16.7%) and, hierarchically, for infections by HPV33, 16, 35, 59, 31 and 52 (in decreasing order). Referral of women positive for either p16 or LSIL+ cytology had 97.8% sensitivity for CIN2+ and women negative for both of these had a 3-year CIN3+ risk of 0.2%. Similar results were seen for women being either p16 or HPV16/33 positive. hrHPV-positive women who were negative for p16 and cytology (LSIL threshold) had a very low CIN3+ rate in the following 3 years. Recalling them after that interval and referring those positive for either test to immediate colposcopy seem to be an efficient triage strategy. The same applies to p16 and HPV16.     

Considerations for Measurement of Embryonic Organ Growth

Organogenesis is a complex coordinated process of cell proliferation, growth, migration, and apoptosis. Differential growth rates, particularly during cardiogenesis, play a role in establishing morphology. Studies using stereological and cell sorting methods derive averages of morphogenetic parameters for an organ. To understand tissue composition and differential growth, the researcher must determine a number of morphogenetic parameters in the developing organ. Such measurements require sectioning to enable identification of organ borders, tissue components and cell types, three-dimensional (3D)-reconstruction of sections to visualize morphology and a 3D-measurement scheme to build local morphogenetic information. Although thick the section confocal microscopy partially solves these issues, information loss at the section surface hampers the reconstruction of 3D morphology. Episcopic imaging provides the correct morphology but lacks histological procedures to identify multiple cell types. The 3D-measurement scheme is based on systematic sampling, with overlapping sample volumes, of the entire organ in the aligned image stack. For each sample volume, morphogenetic variables are calculated and results projected back to the cube (boxel) at the sample volume center. Boxel size determines spatial resolution of the final quantitative 3D-reconstruction whereas size of the sample volume determines the precision of the morphogenetic information. The methods described here can be used to measure tissue volume, proliferation and cell size, to determine contribution and distribution of cell types in a tissue and to display this information in a quantitative 3D-reconstruction. Anat Rec, 302:49-57, 2019. © 2018 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists     

IgG4‐related disease of the paratestis with the scrotal fluid: A case report

IgG4‐related disease (IgG4‐RD) can affect various organs, and the pancreas and salivary gland are representative examples. We report a rare case of IgG4‐RD of the paratestis. A 74‐year‐old man presented with left scrotal swelling. Scrotopuncture drainage and cytology confirmed a clear, yellow retention liquid (130 mL) with many small, similar lymphocytes and a few plasmacytes. Many lymphoid cells were immunopositive for CD3 on a cell block section, indicating that a predominant type of lymphoid cells was T cell. There were also some CD20 immunopositive cells and a few IgG4 immunopositive cells. Two months later the left scrotal swelling had returned, and he underwent radical inguinal orchiectomy. Microscopically, there was considerable lymphoplasmacytic inflammatory infiltration, fibrosis and abundant IgG4 immunopositive cells in the paratesticular region. The histopathologic and immunohistochemistry findings were consistent with IgG4‐RD. However, the abundant T cells in the scrotal fluid complicated the cytological diagnosis in our case.     

Cytokeratin phenotypes at the dento-gingival junction in relative health and inflammation, in smokers and non-smokers

OBJECTIVES: The cells of the junctional epithelium (JE) provide and maintain the epithelial attachment, and remain morphologically and phenotypically distinct from oral sulcular (OSE) and external oral epithelia (EOE), from which they may be regenerated de novo. Expression of cytokeratins (CK) in human epithelia has been shown to be highly site-specific, implying a functional role. The aims of this study were to differentiate between the cyto-keratin profiles of JE, OSE, EOE and pocket epithelia (PE) in health and disease, in smokers and non-smokers.
MATERIALS AND METHODS: The cytokeratin profiles of 40 samples of healthy and clinically inflamed human gingival tissue taken from 15 smokers and 25 non-smokers were studied by immunocytochemistry. Cryostat sections of fresh frozen gingival tissues were stained with a panel of monoclonal antibodies (mAb) and visualised by a biotin-Streptavidin-peroxidase complex technique.
RESULTS: JE and PE expressed an identical range of cytokeratins irrespective of the inflammatory or smoking status, with the exception of CK4 expression, which tended to be increased in smokers. The OSE and EOE expressed non-cornifying and cornifying differentiation cytokeratins respectively, but in the presence of inflammation, both these epithelia showed increased expression of CK19 at a basal level in association with expression of one or more of the simple cytokeratins. JE/PE expressed CK17 in external layers only, approximating the tooth surface. All epithelia expressed CK6,16 the markers of high cell turnover.
CONCLUSIONS: CK19 was a consistent differentiation marker for JE and PE.Expression of CK8,18 was enhanced by inflammation. CK4 expression increased in association with smoking. Markers of differentiation were not always co-expressed equally within a pair. Pairs were not always completely mutually exclusive with frequent co-localisation.     

PTTG和bFGF在舌癌细胞株Tca8113中表达相关性研究

目的：探讨垂体肿瘤转化基因（Pituitary Tumor Transforming Gene,PTrG）蛋白在舌癌细胞株Tca8113中的表达及其与碱性成纤维细胞生长因子（basic Fibroblast Growth Factor,bFGF）间的关系.方法：应用免疫细胞化学方法,检测加入bFGF抗体后孵育的癌细胞中PTTG蛋白表达的变化.结果：Tca8113中PTTG蛋白呈阳性表达,并受bFGF影响,加入bFGF抗体的培养液中的癌细胞PTTG蛋白荧光明显减弱,随着时间延长和bFGF抗体滴度增加,减弱趋势明显.结论：舌癌细胞株中PTTG蛋白的表达受bFGF制约,对二者的联合治疗可能是一种治疗舌癌的有效方法.     

Characterization of antigen-presenting cells in human apical periodontitis lesions by flow cytometry and immunocytochemistry

AIM: To analyse phenotypic characteristics of antigen-presenting cells (APC), isolated from human periapical lesions by flow cytometry and immunocytochemistry. METHODOLOGY: Sixteen periapical lesions were digested for 15 min with 0.05% collagenase. Mononuclear cells, separated from other inflammatory cells by density centrifugation, were processed for flow cytometry and/or immunocytochemistry. Single and double immunostainings were performed using monoclonal antibodies specific for human CD45, CD3, CD19, CD14, HLA-DR, CD1a, CD83 and CD123. RESULTS: Antigen-presenting cells (HLA-DR(+) cells) represented 32.9 +/- 17.8% of total mononuclear cells. Amongst them, B cells (HLA-DR(+) CD19(+)) were the predominant APC population, followed by activated macrophages (HLA-DR(+) CD14(+)), dendritic cells (DC) (HLA-DR(+) CD14(-) CD19(-) CD3(-)) and activated T cells (HLA-DR(+) CD3(+)). Based on the predominance of T cells (CD3(+)) or B cells and plasma cells (CD19(+) and CD19(lo), respectively) amongst mononuclear cell infiltrates, lesions were divided into T- and B-types. The percentage of DC in T-type lesions (27.1 +/- 6.8% of total HLA-DR(+) cells) was higher, compared with B-type lesions (10.3 +/- 5.2%) (P < 0.01). Within the DC population, the percentages of CD1a (Langerhans cell type) and CD123 (probably plasmacytoid DC type) did not differ significantly between the groups (P > 0.05). However, the percentage of mature DC (CD83(+)) was significantly higher in T-type periapical lesions (P < 0.05). CONCLUSIONS: Flow cytometry and immunocytochemistry are suitable methods for phenotypic analysis of APC after their isolation from human periapical lesions. APC, that were phenotypically heterogeneous, constituted a significant component of infiltrating cells. Lesions with the predominance of T cells were characterized by a higher proportion of mature DC (HLA-DR(+)CD83(+) cells) than lesions with predominance of B cells/plasma cells.     

Differential localization of laminin gamma and integrin beta in primary cultures of the rat gingival epithelium

OBJECTIVES: The aim of this study was to investigate the differential immunolocalization of laminin gamma(2) and integrin beta(4) in primary cultures of the rat gingival epithelium. METHODS: The gingival epithelium was obtained from Sprague-Dawley rats and was cultured in serum-free keratinocyte growth medium (DK-SFM). Western blotting analysis, immunofluorescence, confocal laser scanning microscopy (CLSM), and immuno-gold labeling for laminin gamma(2) and integrin beta(4) were employed. CLSM images for laminin and integrin were analyzed in horizontal (x-y axis) and in vertical (x-z axis) sections. RESULTS: Both laminin gamma(2) and integrin beta(4) were detected by Western blot analysis in the gingival epithelium. Immunolocalization of laminin gamma(2) was distinct in the cytoplasm to form one or two irregular rings in gingival epithelial cells. By contrast, integrin beta(4) was localized diffusely in the cytoplasm. F-actin (indicating actin filaments) was clearly discernible at the periphery of the cytoplasm to form a cellular fringe. In x-z axis images obtained by CLSM, laminin gamma(2) was recognized as large foci in the most inner portion just above the basal plasma membrane. Integrin beta(4) existed in the area where F-actin was labeled surrounding the membrane. Immuno-electron microscopy showed that 10nm colloidal gold particles indicating laminin gamma(2) were mainly localized at the extracellular portion and in the peripheral cytoplasm, whereas integrin beta(4) was distributed in the cytoplasm close to the basal plasma membrane but not in extracellular regions. CONCLUSIONS: In primary cultures of the rat gingival epithelium, both laminin gamma(2) and integrin beta(4) may be produced by the epithelium, and irregular rings of laminin gamma(2) are formed in areas where gingival cells adhere to the extracellular matrix.     

The cholinergic basal forebrain in the ferret and its inputs to the auditory cortex

Cholinergic inputs to the auditory cortex can modulate sensory processing and regulate stimulus‐specific plasticity according to the behavioural state of the subject. In order to understand how acetylcholine achieves this, it is essential to elucidate the circuitry by which cholinergic inputs influence the cortex. In this study, we described the distribution of cholinergic neurons in the basal forebrain and their inputs to the auditory cortex of the ferret, a species used increasingly in studies of auditory learning and plasticity. Cholinergic neurons in the basal forebrain, visualized by choline acetyltransferase and p75 neurotrophin receptor immunocytochemistry, were distributed through the medial septum, diagonal band of Broca, and nucleus basalis magnocellularis. Epipial tracer deposits and injections of the immunotoxin ME20.4‐SAP (monoclonal antibody specific for the p75 neurotrophin receptor conjugated to saporin) in the auditory cortex showed that cholinergic inputs originate almost exclusively in the ipsilateral nucleus basalis. Moreover, tracer injections in the nucleus basalis revealed a pattern of labelled fibres and terminal fields that resembled acetylcholinesterase fibre staining in the auditory cortex, with the heaviest labelling in layers II/III and in the infragranular layers. Labelled fibres with small en‐passant varicosities and simple terminal swellings were observed throughout all auditory cortical regions. The widespread distribution of cholinergic inputs from the nucleus basalis to both primary and higher level areas of the auditory cortex suggests that acetylcholine is likely to be involved in modulating many aspects of auditory processing.     

人胎肝干细胞体外分离、培养及鉴定

研究人胎肝干细胞体外分离纯化方法，并对其进行初步鉴定及生物学特性分析。采用胶原酶消化、重力沉降及密度梯度离心方法分离人胎肝干细胞，通过免疫细胞化学方法对其进行初步鉴定，以及应用流式细胞仪等对其生长状况进行评估。所获得的原代细胞共传8代，细胞呈小梭形或三角形，体积较小，胞核较大，而胞浆较少；第4代细胞中进入生长期的细胞约占88．2％；免疫细胞化学染色显示细胞胞质中ALB、CK19染色阳性。人胎肝中存在具有肝干细胞特征的细胞，它们可能会成为细胞移植及肝移植良好的细胞资源。