Is human chorionic gonadotropin directly involved in the regulation of human implantation?

The regulation of human implantation is not fully understood. hCG as one of the earliest embryonal signals may be a major regulator in the parakrine embryo-endometrial communication. The expression of full-length hCG/LH-receptor mRNA could be demonstrated in human endometrium throughout the follicular and secretory phase of the menstrual cycle. In contrast, in early pregnancy decidua only truncated variants could be detected. To investigate direct effects of hCG on the human endometrium, an intrauterine microdialysis device was developed to measure parakrine mediators within the uterine cavity in vivo. Using this system, hCG was applied in the secretory phase and the endometrial response was evaluated. The administration of hCG (500 IU/ml) provoked a significant inhibition of intrauterine IGFBP-1 and M-CSF, while LIF, VEGF and MMP-9 were significantly stimulated. Taken together there appear to be multiple direct effects of hCG on the endometrium that precede the classical endocrine role of the hormone.     

Genetic polymorphism and human cancer

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Modulation of human embryonic stem cell-derived cardiomyocyte growth: a testbed for studying human cardiac hypertrophy?

Human embryonic stem cell-derived cardiomyocytes (hESC-CM) are being developed for tissue repair and as a model system for cardiac physiology and pathophysiology. However, the signaling requirements of their growth have not yet been fully characterized. We showed that hESC-CM retain their capacity for increase in size in long-term culture. Exposing hESC-CM to hypertrophic stimuli such as equiaxial cyclic stretch, angiotensin II, and phenylephrine (PE) increased cell size and volume, percentage of hESC-CM with organized sarcomeres, levels of ANF, and cytoskeletal assembly. PE effects on cell size were separable from those on cell cycle. Changes in cell size by PE were completely inhibited by p38-MAPK, calcineurin/FKBP, and mTOR blockers. p38-MAPK and calcineurin were also implicated in basal cell growth. Inhibitors of ERK, JNK, and CaMK II partially reduced PE effects; PKG or GSK3β inhibitors had no effect. The role of p38-MAPK was confirmed by an additional pharmacological inhibitor and adenoviral infection of hESC-CM with a dominant-inhibitory form of p38-MAPK. Infection of hESC-CM with constitutively active upstream MAP2K3b resulted in an increased cell size, sarcomere and cytoskeletal assembly, elongation of the cells, and induction of ANF mRNA levels. siRNA knockdown of p38-MAPK inhibited PE-induced effects on cell size. These results reveal an important role for active protein kinase signaling in hESC-CM growth and hypertrophy, with potential implications for hESC-CM as a novel in vitro test system. This article is part of a special issue entitled, "Cardiovascular Stem Cells Revisited".     

β-Mannosidase in human leukocytes and fibroblasts

In human leukocytes and fibroblasts -mannosidase activity has a unimodal pH optimum (4.0–4.5). Markedly reduced activity is found in I-cell disease. Normal activities in human fibroblasts are ten times higher than in the goat, in which species a deficiency disease has been described.     

Can we get human nature right?

Few questions in science are as controversial as human nature. At stake is whether our basic concepts and emotions are all learned from experience, or whether some are innate. Here, I demonstrate that reasoning about innateness is biased by the basic workings of the human mind. Psychological science suggests that newborns possess core concepts of “object” and “number.” Laypeople, however, believe that newborns are devoid of such notions but that they can recognize emotions. Moreover, people presume that concepts are learned, whereas emotions (along with sensations and actions) are innate. I trace these beliefs to two tacit psychological principles: intuitive dualism and essentialism. Essentialism guides tacit reasoning about biological inheritance and suggests that innate traits reside in the body; per intuitive dualism, however, the mind seems ethereal, distinct from the body. It thus follows that, in our intuitive psychology, concepts (which people falsely consider as disembodied) must be learned, whereas emotions, sensations, and emotions (which are considered embodied) are likely innate; these predictions are in line with the experimental results. These conclusions do not speak to the question of whether concepts and emotions are innate, but they suggest caution in its scientific evaluation.     

How reliable are human phylogenetic hypotheses?

Cladistic analysis of cranial and dental evidence has been widely used to generate phylogenetic hypotheses about humans and their fossil relatives. However, the reliability of these hypotheses has never been subjected to external validation. To rectify this, we applied identical methods to equivalent evidence from two groups of extant higher primates for whom reliable molecular phylogenies are available, the hominoids and papionins. We found that the phylogenetic hypotheses based on the craniodental data were incompatible with the molecular phylogenies for the groups. Given the robustness of the molecular phylogenies, these results indicate that little confidence can be placed in phylogenies generated solely from higher primate craniodental evidence. The corollary of this is that existing phylogenetic hypotheses about human evolution are unlikely to be reliable. Accordingly, new approaches are required to address the problem of hominin phylogeny.     

Immunosurveillance function of human mast cell？

Mast cell (MC) is so widely recognized as a critical effector in allergic disorders that it can be difficult to think of MC in any other context. Indeed, MCs are multifunctional and recently shown that MCs can also act as antigen presenters as well as effector elements of human immune system. First observations of their possible role as anti-tumor cells in peri- or intra-tumoral tissue were mentioned five decades ago and a high content of MCs is considered as a favorable prognosis, consistent with this study. Believers of this hypothesis assumed them to be inhibitors of tumor development through their pro-apoptotic and -necrolytic granules e.g., granzymes and TNF-α. However, some still postulate them to be enhancers of tumor development through their effects on angiogenesis due to mostly tryptase. There are also some data suggesting increased MC density causes tumor development and indicates bad prognosis. Furthermore, since MC-associated mediators have shown to influence various aspects of tumor biology, the net effect of MCs on the development/ progression of tumors has been difficult to evaluate. For instance, chymase induces apoptosis in targets; yet, tryptase, another MC protease, is a well-known mitogen. MCs with these various enzyme expression patterns may mediate different functions and the predominant MC type in tissues may be determined by the environmental needs. The coexistence of tryptase-expressing MCs (MCT) and chymase and tryptase-expressing MCs (MCTC) in physiological conditions reflects a naturally occurring balance that contributes to tissue homeostasis. We have recently discussed the role and relevance of MC serine proteases in different bone marrow diseases.     

Structural basis of human γ-secretase assembly

The four-component intramembrane protease γ-secretase is intricately linked to the development of Alzheimer’s disease. Despite recent structural advances, the transmembrane segments (TMs) of γ-secretase remain to be specifically assigned. Here we report a 3D structure of human γ-secretase at 4.32-Å resolution, determined by single-particle, electron cryomicroscopy in the presence of digitonin and with a T4 lysozyme fused to the amino terminus of presenilin 1 (PS1). The overall structure of this human γ-secretase is very similar to that of wild-type γ-secretase determined in the presence of amphipols. The 20 TMs are unambiguously assigned to the four components, revealing principles of subunit assembly. Within the transmembrane region, PS1 is centrally located, with its amino-terminal fragment (NTF) packing against Pen-2 and its carboxyl-terminal fragment (CTF) interacting with Aph-1. The only TM of nicastrin associates with Aph-1 at the thick end of the TM horseshoe, and the extracellular domain of nicastrin directly binds Pen-2 at the thin end. TM6 and TM7 in PS1, which harbor the catalytic aspartate residues, are located on the convex side of the TM horseshoe. This structure serves as an important framework for understanding the function and mechanism of γ-secretase.Alzheimer’s disease (AD), characterized by formation of β-amyloid plaque in the brain of a patient, is closely associated with γ-secretase (1, 2). Amyloid precursor protein (APP) is processed by β-secretase in the extracellular space to produce a membrane-tethered fragment known as C99 (3). APP C99 then undergoes sequential cleavages by γ-secretase, generating a series of β-amyloid peptides (Aβ) exemplified by Aβ42 and Aβ40 (4, 5). Among all Aβs, Aβ42 is particularly prone to aggregation, resulting in formation of β-amyloid plaque and presumably contributing to the development of AD (6).Mature γ-secretase contains four components: presenilin, Pen-2, nicastrin, and Aph-1. The catalytic subunit presenilin is predicted to contain nine transmembrane segments (TMs), with two catalytic aspartate residues on TM6 and TM7. During assembly of γ-secretase, presenilin undergoes an autocatalytic cleavage to yield two polypeptide fragments, NTF (comprising TMs 1–6) and CTF (comprising TMs 7–9) (7, 8). PS1 is the target of most mutations derived from early onset familial Alzheimer’s disease patients (1). The largest component nicastrin has only one TM but contains a highly glycosylated extracellular domain (ECD), which presumably recognizes the amino terminus of substrate protein (9–11). The smallest component Pen-2 is thought to be required for the autocatalytic maturation of presenilin and γ-secretase activity (12, 13). Aph-1, required for assembly of γ-secretase (14), appears to have a previously unidentified fold with seven predicted TMs.The assembly and intersubunit interactions of γ-secretase constitute an important basis for its mechanistic understanding and have been extensively investigated during the past decade. As the central component of γ-secretase, PS1 was shown to interact with both Pen-2 and Aph-1 and form distinct subcomplexes (15–21). The only TM of nicastrin was thought to bind Aph-1 and contribute to interactions with PS1. Rationalization of these biochemical findings and other functional observations requires detailed 3D structural information on γ-secretase.In contrast to rapid accumulation of biochemical and functional data on γ-secretase, structural determination has been slow to emerge, largely due to the technical challenges associated with expression and manipulation of the intact γ-secretase. Several EM analyses have yielded low-resolution images of γ-secretase (22–27), with the overall shapes diverging from each other. Investigation of γ-secretase by other biophysical methods produced an NMR structure of the presenilin CTF (28) and X-ray structures of an archaeal homolog of presenilin (29) and a eukaryotic homolog of nicastrin (30).The high-resolution cryo-electron microscopy (cryo-EM) structure of human γ-secretase, determined at 4.5-Å resolution and in the presence of amphipols, revealed an overall architecture that is qualitatively different from all previous structures (31). The EM densities allowed identification of 19 TMs and construction of an atomic model for the ECD (31). However, these densities lacked connectivity between TMs and exhibited few side-chain features in the TMs, disallowing specific TM assignment to the four components. The use of amphipols also raises the question of whether the structure of human γ-secretase is dependent upon the choice of detergent used. In this study, we address these concerns and report, to our knowledge, the first structure of an intact γ-secretase with all TMs assigned.     

Filamentous-actins in human hepatocarcinoma cells with CLSM

AIM:To establish a method for optical sections of HepG2human hepatoblastoma cells with confocal laser scanningmicroscope (CLSM) and to study the spatial structure offilamentous actin (F-actin) in HepG2 cells.METHODS:HepG2 cells were stained with FITC-phalloidinthat specifically binds F-actin,with propidium iodide (PI) tothe nucleus,and scanned with a CLSM to generate opticallysectioned images.A series of optical sections takensuccessively at different focal levels in steps of 0.7μm werereconstructed with the CLSM reconstruction program.RESULTS:CLSM images showed that the FITC-stained F-actin was abundant microfilament bundles parallel or nettedthrough the whole cell and its processes.Most F-actinmicrofilaments extended through the cell from one part towardthe other or run through the process.Some microfilamentswere attached to the plasma membrane,or formed astructural bridge connecting to the neighboring cells.CONCLUSION:A method for double labeling HepG2 humanhepatoblastoma cells and CLSM imaging F-actin microfilamentsand nuclei by image thin optical sections and spatial structurewas developed.It provides a very useful way to study thespatial structure of F-actin.     

Treatment of human hepatocellular carcinoma by fibroblast-mediated human interferon α gene therapy in combination with adoptive chemoimmunotherapy

The therapeutic effect of the fibroblast-mediated human interferon (IFN) gene therapy in combination with interleukin-2 (IL-2) activated killer cells (AK)/doxorubicin (i.e., adoptive chemoimmunotherapy) on nude mice bearing the human hepatocellular carcinoma (HCC) was investigated. A fibroblast cell clone (NIH3T3-IFN⁺) secreting 1024 U/ml human IFN was obtained from 14 positive clones by BMGNeo-INF DNA transfection, G418-resistant selection, limiting dilution and assay of IFN activity. After i.p. implantation of NIH3T3-IFN⁺ encapsulated into collagen, serum human IFN activity could be detected from 12 h to day 15 with a peak at 72 h. AK were prepared from human peripheral mononuclear cells costimulated in vitro by IL-2 and inactivated human SMMC 7721 HCC cells. When the NIH3T3-IFN⁺ cells were i.p. implanted into the HCC-bearing nude mice, the grown of HCC was inhibited and the survival time of the mice was extended. The growth of HCC was inhibited more obviously when AK was i.v. injected and IL-2 was i.p. injected after the NIH3T3-IFN⁺ cells had been implanted. The best therapeutic effect was achieved when NIH3T3-IFN⁺ cells were used in combination with IL-2/AK/doxorubicin. All these results suggested that the fibroblast-mediated human IFN gene therapy could be used to treat the human hepatocellular carcinoma effectively and that when used in combination with IL-2-based adoptive chemoimmunotherapy, the therapeutic effect would be better.Abbreviations IFN interferon - HCC hepatocellular carcinoma - IL-2 interleukin-2 - AK activated killer cells - Dox doxorubicin This research was supported by the National Natural Sciences Foundation of China (grant 39421009)