Tissue culture of human thyroid cancer

Human thyroid cancer cells in the pleural effusion were serially cultivated in vitro. Three kinds of cell lines were established from the same primary culture and were designated as PS, TS and TR lines, respectively. These three have been cultured for 574 days up to May 1, 1968. The cells of PS and TR lines were epithelial-like, whereas those of TS line revealed fibroblastic character. The chromosome numbers of PS and TR lines exhibited the modes near the hypertetraploid region, while TS line showed the mode of hypo-triploid number. Eosinophilic particles which were stained metachromatically by toluidine blue were present in the cytoplasm of these cells. The histochemical findings of the cells of each line were identical with those of thyroid cancer cells in vivo. The cells aggregated by the gyratory culture showed epithelial characters under microscopic observation of the sectioned specimens. The tumors produced in conditoned hamsters demonstrated undifferentiated cancer, which resembled the metastatic thyroid cancer of the patient. Neither collagen nor argentaffine fibers were detected with Van Gieson staining or silver impregenation.</p

Infection of human cytomegalovirus in cultured human gingival tissue.

BackgroundHuman cytomegalovirus (HCMV) infection in the oral cavity plays an important role in its horizontal transmission and in causing viral-associated oral diseases such as gingivitis. However, little is currently known about HCMV pathogenesis in oral mucosa, partially because HCMV infection is primarily limited to human cells and few cultured tissue or animal models are available for studying HCMV infection.ResultsIn this report, we studied the infection of HCMV in a cultured gingival tissue model (EpiGingival, MatTek Co.) and investigated whether the cultured tissue can be used to study HCMV infection in the oral mucosa. HCMV replicated in tissues that were infected through the apical surface, achieving a titer of at least 300-fold at 10 days postinfection. Moreover, the virus spread from the apical surface to the basal region and reduced the thickness of the stratum coreum at the apical region. Viral proteins IE1, UL44, and UL99 were expressed in infected tissues, a characteristic of HCMV lytic replication in vivo. Studies of a collection of eight viral mutants provide the first direct evidence that a mutant with a deletion of open reading frame US18 is deficient in growth in the tissues, suggesting that HCMV encodes specific determinants for its infection in oral mucosa. Treatment by ganciclovir abolished viral growth in the infected tissues.ConclusionThese results suggest that the cultured gingival mucosa can be used as a tissue model for studying HCMV infection and for screening antivirals to block viral replication and transmission in the oral cavity

Stepped vitrification technique for human ovarian tissue cryopreservation

The advantage of stepped vitrification (SV) is avoiding ice crystal nucleation, while decreasing the toxic effects of high cryoprotectant concentrations. We aimed to test this method for human ovarian tissue cryopreservation. Ovarian cortex was taken from 7 fertile adult women. Samples were subjected to an SV protocol performed in an automatic freezer, which allowed sample transfer to ever higher concentrations of dimethyl sulfoxide (DMSO) as the temperature was reduced. Histological evaluation of the vitrified-warmed tissue showed large numbers of degenerated follicles after 24 hours of in vitro culture. We therefore evaluated DMSO perfusion rates by X-ray computed tomography, ice crystal formation by freeze-substitution, and cell toxicity by transmission electron microscopy, seeking possible reasons why follicles degenerated. Although cryoprotectant perfusion was considered normal and no ice crystals were formed in the tissue, ultrastructural analysis detected typical signs of DMSO toxicity, such as mitochondria degeneration, alterations in chromatin condensation, cell vacuolization and extracellular matrix swelling in both stromal and follicular cells. The findings indicated that the method failed to preserve follicles due to the high concentrations of DMSO used. However, adaptations can be made to avoid toxicity to follicles caused by elevated levels of cryoprotectants.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) 2016/22947-

Machine learning classification of human joint tissue from diffuse reflectance spectroscopy data

Objective: To assess if incorporation of DRS sensing into real-time robotic surgery systems has merit. DRS as a technology is relatively simple, cost-effective and provides a non-contact approach to tissue differentiation. Methods: Supervised machine learning analysis of diffuse reflectance spectra was performed to classify human joint tissue that was collected from surgical procedures. Results: We have used supervised machine learning in the classification of a DRS human joint tissue data set and achieved classification accuracy in excess of 99%. Sensitivity for the various classes were; cartilage 99.7%, subchondral 99.2%, meniscus 100% and cancellous 100%. Full wavelength range is required for maximum classification accuracy. The wavelength resolution must be larger than 8nm. A SNR better than 10:1 was required to achieve a classification accuracy greater than 50%. The 800-900nm wavelength range gave the greatest accuracy amongst those investigated. Conclusion: DRS is a viable method for differentiating human joint tissue and has the potential to be incorporated into robotic orthopaedic surgery

A microfluidic chip based model for the study of full thickness human intestinal tissue using dual flow

© 2016 Author(s). The study of inflammatory bowel disease, including Ulcerative Colitis and Crohn's Disease, has relied largely upon the use of animal or cell culture models; neither of which can represent all aspects of the human pathophysiology. Presented herein is a dual flow microfluidic device which holds full thickness human intestinal tissue in a known orientation. The luminal and serosal sides are independently perfused ex vivo with nutrients with simultaneous waste removal for up to 72 h. The microfluidic device maintains the viability and integrity of the tissue as demonstrated through Haematoxylin & Eosin staining, immunohistochemistry and release of lactate dehydrogenase. In addition, the inflammatory state remains in the tissue after perfusion on the device as determined by measuring calprotectin levels. It is anticipated that this human model will be extremely useful for studying the biology and tes ting novel interventions in diseased tissue

α_1L-adrenoceptors mediate contraction of human erectile tissue

α1-adrenoceptor antagonists can impact upon sexual function and have potential in the treatment of erectile dysfunction. Human erectile tissue contains predominantly α1A-adrenoceptors, and here we examined whether contractions of this tissue are mediated by the functional phenotype, the α1L-adrenoceptor. Functional experiments using subtype selective agonists and antagonists, along with radioligand ([3H]tamsulosin) binding assays, were used to determine the α1-adrenoceptor population. A61603, a α1A-adrenoceptor agonist, was a full agonist with a potency 21-fold greater than that of noradrenaline. The α1A- and α1D-adrenoceptor antagonist tamsulosin antagonized noradrenaline responses with high affinity (pKD = 9.7 ± 0.3), whilst BMY7378 (100 nM) (α1D-adrenoceptor antagonist) failed to antagonize responses. In contrast, relatively low affinity estimates were obtained for both prazosin (pKD = 8.2 ± 0.1) and RS17053 (pKD = 6.9 ± 0.2), antagonists which discriminate between the α1A- and α1L-adrenoceptors. [3H]Tamsulosin bound with high affinity to the receptors of human erectile tissue (pKD = 10.3 ± 0.1) with a receptor density of 28.1 ± 1.4 fmol mg−1 protein. Prazosin displacement of [3H]tamsulosin binding revealed a single homogenous population of binding sites with a relatively low affinity for prazosin (pKi = 8.9). Taken together these data confirm that the receptor mediating contraction in human erectile tissue has the pharmacological properties of the α1L-adrenoceptor. Keywords: Erectile tissue, α1-adrenoceptor subtypes, α1L-adrenoceptor, Tamsulosin, Prazosi

Large-scale discovery of enhancers from human heart tissue.

Development and function of the human heart depend on the dynamic control of tissue-specific gene expression by distant-acting transcriptional enhancers. To generate an accurate genome-wide map of human heart enhancers, we used an epigenomic enhancer discovery approach and identified ∼6,200 candidate enhancer sequences directly from fetal and adult human heart tissue. Consistent with their predicted function, these elements were markedly enriched near genes implicated in heart development, function and disease. To further validate their in vivo enhancer activity, we tested 65 of these human sequences in a transgenic mouse enhancer assay and observed that 43 (66%) drove reproducible reporter gene expression in the heart. These results support the discovery of a genome-wide set of noncoding sequences highly enriched in human heart enhancers that is likely to facilitate downstream studies of the role of enhancers in development and pathological conditions of the heart