Identification of Chicken CD44 as a Novel B Lymphocyte Receptor for Infectious Bursal Disease Virus.

Infectious bursal disease virus (IBDV), which targets bursa B lymphocytes, causes severe immunosuppressive disease in chickens, inducing huge economic losses for the poultry industry. To date, the functional receptor for IBDV binding and entry into host cells remains unclear. This study used mass spectrometry to screen host proteins of chicken bursal lymphocytes interacting with VP2. The chicken transmembrane protein cluster of differentiation 44 (chCD44) was identified and evaluated for its interaction with IBDV VP2, the major capsid protein. Overexpression and knockdown experiments showed that chCD44 promotes replication of IBDV. Furthermore, soluble chCD44 and the anti-chCD44 antibody blocked virus binding. The results of receptor reconstitution indicated that chCD44 overexpression conferred viral binding capability in nonpermissive cells. More important, although we found that IBDV could not replicate in the chCD44-overexpressed nonpermissive cells, the virus could enter nonpermissive cells using chCD44. Our finding reveals that chCD44 is a cellular receptor for IBDV, facilitating virus binding and entry in target cells by interacting with the IBDV VP2 protein. IMPORTANCE Infectious bursal disease virus (IBDV) causes severe immunosuppressive disease in chickens, inducing huge economic losses for the poultry industry. However, the specific mechanism of IBDV invading host cells of IBDV was not very clear. This study shed light on which cellular protein component IBDV is used to bind and/or enter B lymphocytes. The results of our study revealed that chCD44 could promote both the binding and entry ability of IBDV in B lymphocytes, acting as a cellular receptor for IBDV. Besides, this is the first report about chicken CD44 function in viral replication. Our study impacts the understanding of the IBDV binding and entry process and sets the stage for further elucidation of the infection mechanism of IBDV.

Lactoferrin-Hexon Interactions Mediate CAR-Independent Adenovirus Infection of Human Respiratory Cells.

Virus entry into host cells is a complex process that is largely regulated by access to specific cellular receptors. Human adenoviruses (HAdVs) and many other viruses use cell adhesion molecules such as the coxsackievirus and adenovirus receptor (CAR) for attachment to and entry into target cells. These molecules are rarely expressed on the apical side of polarized epithelial cells, which raises the question of how adenoviruses-and other viruses that engage cell adhesion molecules-enter polarized cells from the apical side to initiate infection. We have previously shown that species C HAdVs utilize lactoferrin-a common innate immune component secreted to respiratory mucosa-for infection via unknown mechanisms. Using a series of biochemical, cellular, and molecular biology approaches, we mapped this effect to the proteolytically cleavable, positively charged, N-terminal 49 residues of human lactoferrin (hLF) known as human lactoferricin (hLfcin). Lactoferricin (Lfcin) binds to the hexon protein on the viral capsid and anchors the virus to an unknown receptor structure of target cells, resulting in infection. These findings suggest that HAdVs use distinct cell entry mechanisms at different stages of infection. To initiate infection, entry is likely to occur at the apical side of polarized epithelial cells, largely by means of hLF and hLfcin bridging HAdV capsids via hexons to as-yet-unknown receptors; when infection is established, progeny virions released from the basolateral side enter neighboring cells by means of hLF/hLfcin and CAR in parallel.IMPORTANCE Many viruses enter target cells using cell adhesion molecules as receptors. Paradoxically, these molecules are abundant on the lateral and basolateral side of intact, polarized, epithelial target cells, but absent on the apical side that must be penetrated by incoming viruses to initiate infection. Our study provides a model whereby viruses use different mechanisms to infect polarized epithelial cells depending on which side of the cell-apical or lateral/basolateral-is attacked. This study may also be useful to understand the biology of other viruses that use cell adhesion molecules as receptors.

Role of PDGF receptor-α during human cytomegalovirus entry into fibroblasts.

Human CMV (HCMV) exhibits a broad cell tropism that depends on two virion glycoprotein complexes: a trimeric complex (gH/gL/gO) that facilitates viral infection primarily in fibroblasts and a pentameric complex (gH/gL/pUL128-pUL130-pUL131A) that mediates infection in epithelial and endothelial cells. We performed genome-wide CRISPR screens in which the PDGF receptor-α (PDGFRα) was identified as the most significant cellular gene product essential for infection by HCMV virions containing only trimeric complex (trimer-only virus). Trimer-only virus did not enter PDGFRα knockout fibroblasts. By using knockout fibroblasts, the extracellular domain of PDGFRα required for virus entry was mapped, and the intracellular tyrosine kinase domain was shown to be nonessential. In addition, direct cell-to-cell spread of virus from knockout cells transfected with trimer-only viral DNA was blocked, despite the production of infectious virus in the transfected cells. In contrast to trimer-only virus, wild-type HCMV virions containing both trimeric and pentameric complexes entered PDGFRα knockout cells, reinforcing the view that fibroblasts contain a second, independent receptor for the pentameric complex. Importantly, however, wild-type virus entered the knockout fibroblasts at reduced efficiency compared with parental fibroblasts, arguing that the cellular receptor for the virion pentameric complex is limiting or that virions are produced containing different relative amounts of the two glycoprotein complexes. Finally, ectopic expression of PDGFRα in ARPE-19 epithelial cells and THP-1 monocytic cells, which have little to no endogenous PDGFRα expression, markedly enhanced their susceptibility to trimer-only virions. In sum, our data clarify several key determinants of HCMV tropism.

Subcutaneous panniculitis-like T-cell lymphoma in a 14-year-old female homozygous for HAVCR2 mutation.

Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) is a rare cytotoxic T-cell lymphoma preferentially involving subcutis. A link between patients with SPTCL and HAVCR2 mutations has recently been discovered. We present a 14-year-old girl of Chinese heritage who was diagnosed with SPTCL in the context of homozygous HAVCR2 status for c.245A>G p. (Tyr82Cys) and achieved complete remission after treatment with cyclosporin and steroids. Dermatologists should be aware of the diagnostic, management and familial genetic counselling utility of HAVCR2 for investigating and managing patients with SPTCL.

Mechanically Triggered Hybridization Chain Reaction.

Cells transmit piconewton forces to receptors to mediate processes such as migration and immune recognition. A major challenge in quantifying such forces is the sparsity of cell mechanical events. Accordingly, molecular tension is typically quantified with high resolution fluorescence microscopy, which hinders widespread adoption and application. Here, we report a mechanically triggered hybridization chain reaction (mechano-HCR) that allows chemical amplification of mechanical events. The amplification is triggered when a cell receptor mechanically denatures a duplex revealing a cryptic initiator to activate the HCR reaction in situ. Importantly, mechano-HCR enables direct readout of pN forces using a plate reader. We leverage this capability and measured mechano-IC50 for aspirin, Y-27632, and eptifibatide. Given that cell mechanical phenotypes are of clinical importance, mechano-HCR may offer a convenient route for drug discovery, personalized medicine, and disease diagnosis.

Voltage-Dependent Anion Channel Protein 2 (VDAC2) and Receptor of Activated Protein C Kinase 1 (RACK1) Act as Functional Receptors for Lymphocystis Disease Virus Infection.

In previous research, a 27.8-kDa protein in flounder Paralichthys olivaceus gill (FG) cells was identified as a putative cellular receptor (27.8R), which mediated lymphocystis disease virus (LCDV) infection via interaction with a 32-kDa viral attachment protein (VAP) of LCDV, and monoclonal antibodies (MAbs) against 27.8R and 32-kDa VAP were developed. In this study, the 27.8R was identified as voltage-dependent anion channel protein 2 (VDAC2) and receptor of activated protein C kinase 1 (RACK1) of flounder. Recombinant VDAC2 (rVDAC2) and RACK1 (rRACK1) were obtained by prokaryotic expression, and rabbit anti-VDAC2/RACK1 polyclonal antibodies were prepared. The rVDAC2, rRACK1, and 27.8-kDa proteins in FG cells were recognized by anti-27.8R MAbs and anti-VDAC2/RACK1 polyclonal antibodies simultaneously. Preincubation of FG cells with anti-VDAC2/RACK1 polyclonal antibodies significantly decreased the percentages of LCDV-infected cells and LCDV copy numbers, blocked virus infection, and delayed the development of cytopathic effect. The mRNA expressions of VDAC2 and RACK1 in FG cells were upregulated to maximum levels 12 h and 48 h after LCDV infection, respectively. VDAC2/RACK1 knockdown through short interfering RNA (siRNA) significantly reduced VDAC2/RACK1 expression and LCDV copy numbers in FG cells compared with negative controls, while VDAC2/RACK1 expression on LCDV-nonpermissive epithelial papillosum cells (EPCs) conferred susceptibility to LCDV infection, indicating the VDAC2 and RACK1 were sufficient to allow LCDV entry and infection. All these results collectively showed that VDAC2 and RACK1 function as receptors for LCDV entry and infection.IMPORTANCE Lymphocystis disease virus (LCDV) is the causative agent of lymphocystis disease in fish, which has caused huge economic losses to the aquaculture industry worldwide, but the molecular mechanism underlying the LCDV-host interaction remains unclear. Here, the 27.8-kDa putative cellular receptor for LCDV was identified as voltage-dependent anion channel protein 2 (VDAC2) and receptor of activated protein C kinase 1 (RACK1), and our results revealed that VDAC2 and RACK1 expression was sufficient to allow LCDV entry and that they are functional receptors that initiate LCDV infection for the first time, which leads to a better understanding of the molecular mechanism underlying LCDV infection and virus-host interactions.

Tim-3 promotes cell aggressiveness and paclitaxel resistance through NF-κB/STAT3 signalling pathway in breast cancer cells.

OBJECTIVE: Although T-cell immunoglobulin and mucin-domain containing molecule-3 (Tim-3) has been recognized as a promising target for cancer immunotherapy, its exact role in breast cancer has not been fully elucidated. METHODS: Tim-3 gene expression in breast cancer and its prognostic significance were analyzed. Associated mechanisms were then explored in vitro by establishing Tim-3-overexpressing breast cancer cells. RESULTS: In a pooled analysis of The Cancer Genome Atlas (TCGA) database, Tim-3 gene expression levels were significantly higher (P<0.001) in breast cancer tissue, compared with normal tissues. Tim-3 was a prognosis indicator in breast cancer patients [relapse-free survival (RFS), P=0.004; overall survival (OS), P=0.099]. Tim-3 overexpression in Tim-3low breast cancer cells promoted aggressiveness of breast cancer cells, as evidenced by enhanced proliferation, migration, invasion, tight junction deterioration and tumor-associated tubal formation. Tim-3 also enhanced cellular resistance to paclitaxel. Furthermore, Tim-3 exerted its function by activating the NF-κB/STAT3 signalling pathway and by regulating gene expression [cyclin D1 (CCND1), C-Myc, matrix metalloproteinase-1(MMP1), TWIST, vascular endothelial growth factor (VEGF) upregulation, concomitant with E-cadherin downregulation). Lastly, Tim-3 downregulated tight junction-associated molecules zona occludens (ZO)-2, ZO-1 and occludin, which may further facilitate tumor progression. CONCLUSIONS: Tim-3 plays an oncogenic role in breast cancer and may represent a potential target for antitumor therapy.

Porcine Deltacoronavirus Engages the Transmissible Gastroenteritis Virus Functional Receptor Porcine Aminopeptidase N for Infectious Cellular Entry.

Identification of cellular receptors used by coronavirus (CoV) entry into the host cells is critical to an understanding of pathogenesis and to development of intervention strategies. The fourth CoV genus, Deltacoronavirus, evolutionarily related to the Gammacoronavirus, has just been defined recently. In the current study, we demonstrate that porcine aminopeptidase N (pAPN) acts as a cross-genus CoV functional receptor for both enteropathogenic porcine deltacoronovirus (PDCoV) and alphacoronovirus (AlphaCoV) (transmissible gastroenteritis virus [TGEV]) based upon three lines of evidence. First, the soluble S1 protein of PDCoV bound to the surface of target porcine cell lines known to express pAPN as efficiently as TGEV-S1, which could be blocked by soluble pAPN pretreatment. Second, both PDCoV-S1 and TGEV-S1 physically recognized and interacted with pAPN by coimmunoprecipitation in pAPN cDNA-transfected cells and by dot blot hybridization assay. Finally, exogenous expression of pAPN in refractory cells conferred susceptibility to PDCoV-S1 binding and to PDCoV entry and productive infection. PDCoV-S1 appeared to have a lower pAPN-binding affinity and likely consequent lower infection efficiency in pAPN-expressing refractory cells than TGEV-S1, suggesting that there may be differences between these two viruses in the virus-binding regions in pAPN. This study paves the way for dissecting the molecular mechanisms of PDCoV-host interactions and pathogenesis as well as facilitates future vaccine development and intervention strategies against PDCoV infection.IMPORTANCE The emergence of new human and animal coronaviruses is believed to have occurred through interspecies transmission that is mainly mediated by a species-specific receptor of the host. Among the four genera of the Coronavirinae, a couple of functional receptors for the representative members in the genera Alphacoronavirus and Betacoronavirus have been identified, whereas receptors for Gammacoronavirus and Deltacoronavirus, which are believed to originate from birds, are still unknown. Porcine coronaviruses, including the newly discovered porcine deltacoronavirus (PDCoV) associated with diarrhea in newborn piglets, have posed a serious threat to the pork industry in Asia and North America. Here, we report that PDCoV employs the alphacoronavirus TGEV functional receptor porcine aminopeptidase N (pAPN) for cellular entry, demonstrating the usage of pAPN as a cross-genus CoV functional receptor. The identification of the PDCoV receptor provides another example of the expanded host range of CoV and paves the way for further investigation of PDCoV-host interaction and pathogenesis.

HAVCR1 (CD365) and Its Mouse Ortholog Are Functional Hepatitis A Virus (HAV) Cellular Receptors That Mediate HAV Infection.

The hepatitis A virus (HAV) cellular receptor 1 (HAVCR1), classified as CD365, was initially discovered as an HAV cellular receptor using an expression cloning strategy. Due to the lack of HAV receptor-negative replication-competent cells, it was not possible to fully prove that HAVCR1 was a functional HAV receptor. However, biochemistry, classical virology, and epidemiology studies further supported the functional role of HAVCR1 as an HAV receptor. Here, we show that an anti-HAVCR1 monoclonal antibody that protected African green monkey kidney (AGMK) cells against HAV infection only partially protected monkey Vero E6 cells and human hepatoma Huh7 cells, indicating that these two cell lines express alternative yet unidentified HAV receptors. Therefore, we focused our work on AGMK cells to further characterize the function of HAVCR1 as an HAV receptor. Advances in clustered regularly interspaced short palindromic repeat/Cas9 technology allowed us to knock out the monkey ortholog of HAVCR1 in AGMK cells. The resulting AGMK HAVCR1 knockout (KO) cells lost susceptibility to HAV infection, including HAV-free viral particles (vpHAV) and exosomes purified from HAV-infected cells (exo-HAV). Transfection of HAVCR1 cDNA into AGMK HAVCR1 KO cells restored susceptibility to vpHAV and exo-HAV infection. Furthermore, transfection of the mouse ortholog of HAVCR1, mHavcr1, also restored the susceptibility of AGMK HAVCR1 KO cells to HAV infection. Taken together, our data clearly show that HAVCR1 and mHavcr1 are functional HAV receptors that mediate HAV infection. This work paves the way for the identification of alternative HAV receptors to gain a complete understanding of their interplay with HAVCR1 in the cell entry and pathogenic processes of HAV.IMPORTANCE HAVCR1, an HAV receptor, is expressed in different cell types, including regulatory immune cells and antigen-presenting cells. How HAV evades the immune response during a long incubation period of up to 4 weeks and the mechanism by which the subsequent necroinflammatory process clears the infection remain a puzzle that most likely involves the HAV-HAVCR1 interaction. Based on negative data, a recent paper from the S. M. Lemon and W. Maury laboratories (A. Das, A. Hirai-Yuki, O. Gonzalez-Lopez, B. Rhein, S. Moller-Tank, R. Brouillette, L. Hensley, I. Misumi, W. Lovell, J. M. Cullen, J. K. Whitmire, W. Maury, and S. M. Lemon, mBio 8:e00969-17, 2017, https://doi.org/10.1128/mBio.00969-17) suggested that HAVCR1 is not a functional HAV receptor, nor it is it required for HAV infection. However, our data, based on regain of the HAV receptor function in HAVCR1 knockout cells transfected with HAVCR1 cDNA, disagree with their findings. Our positive data show conclusively that HAVCR1 is indeed a functional HAV receptor and lays the ground for the identification of alternative HAV receptors and how they interact with HAVCR1 in cell entry and the pathogenesis of HAV.

SLC10A1 S267F variant influences susceptibility to HBV infection and reduces cholesterol level by impairing bile acid uptake.

The SLC10A1 Ser267Phe (S267F) variant has been reported to severely inhibit hepatitis B virus (HBV) infection and taurocholate transport activity. This study aimed to clarify the effects of this variant on HBV infection and bile acid metabolism. SLC10A1 S267F was genotyped in 2907 HBV-exposed subjects (including HBV persistent carriers and spontaneously recovered subjects) and 1364 unexposed subjects (HBV marker-negative subjects), followed by replication I, comprising 914 exposed subjects and 1123 unexposed subjects, and replication II, comprising 355 children born to HBsAg-positive mothers (226 HBV-infected children and 129 controls). Intriguingly, SLC10A1 AA was observed only in the unexposed group, but not in the exposed group. The SLC10A1 A allele consistently decreased HBV infection risk compared with the G allele (OR = 0.76, 95% CI: 0.64-0.90 in combined samples). In addition, children with the SLC10A1 GA genotype had a reduced risk of perinatal transmission (OR = 0.31, 95% CI: 0.14-0.71). Moreover, unexposed subjects with the SLC10A1 AA genotype exhibited decreased serum total cholesterol and low-density lipoprotein cholesterol compared to those with the GG or GA genotypes (P = 2.975 × 10-4 and 0.004, respectively). The study highlighted the role of the SLC10A1 S267F variant in the loss of the ability to support HBV infection and taurocholate transport activity. Subjects with the AA genotype may escape from HBV infection and present decreased cholesterol levels as a consequence of impaired bile acid uptake.

Cellular entry of white spot syndrome virus and antiviral immunity mediated by cellular receptors in crustaceans.

Enveloped virus usually utilizes the receptor-mediated multiple endocytic routes to enter permissive host cells for successful infection. Cellular receptors are cell surface molecules, either by helping viral attachment to cell surface followed by internalization or by triggering antiviral immunity, participate in the viral-host interaction. White spot syndrome virus (WSSV), the most lethally viral pathogen with envelope and double strand DNA genome in crustacean farming, including shrimp and crayfish, has been recently found to recruit various endocytic routes for cellular entry into host cells. Meanwhile, other than the typical pattern recognition receptors for recognition of WSSV, more and more putative cellular receptors have lately been characterized to facilitate or inhibit WSSV entry. In this review, recent findings on the endocytosis-dependent WSSV entry, viral entry mediated by putative cellular receptors, the molecular interplay between WSSV and cellular receptors, and the following anti-WSSV immunity are summarized and discussed, which may provide us a better understanding of the WSSV pathogenesis and further possible antiviral control of white spot disease in crustacean farming.

Functional characterization of seven-band grouper immunoglobulin like cell adhesion molecule, Nectin4 as a cellular receptor for nervous necrosis virus.

Nectin-4/PVRL4 belonging to the family of immunoglobulin-like cell adhesion molecules was identified as a potential cellular receptor for several animal viruses. Here we show that nervous necrosis virus that causes viral nervous necrosis in teleosts uses the same receptor in its life cycle. Transfection of SSN-1 cell lines with an expression vector encoding Nectin-4 rendered them to be more susceptible to NNV. Immunofluorescence microscopy on Nectin-4 expressing cells revealed that the protein interacted with NNV specifically. A virus binding assay indicated that Nectin-4 was a bonafide receptor that supported virus attachment to the host cell whereas siRNA directed against Nectin-4 blocked NNV infections in grouper primary brain cells. Results of the present study will improve our understanding of the pathogenesis of NNV infection and provide a target for the development of novel antiviral interventions in marine finfish aquaculture.

Increased expression of triggering receptor expressed on myeloid cells 1 and 2 in inflamed human gingiva.

BACKGROUND AND OBJECTIVE: Periodontitis is an infectious disease in which the host immune and inflammatory responses play essential roles in resistance to bacterial infection, as well as the induction of tissue destruction if the immune response is dysregulated. The triggering receptor expressed on myeloid cells (TREMs) modulates inflammatory and innate immune signaling. TREM-1 is considered as an amplifier of the immune response, while TREM-2 is a negative regulator that has yet to be explored in periodontal disease before. We hypothesized that TREMs participated in the innate immune responses during the pathogenesis of periodontitis. Therefore, the aim of this study was to evaluate TREM-1 and TREM-2 expression in the gingival tissues from patients with chronic periodontitis and healthy subjects as well as their correlation with clinical periodontal parameters. This study is the first to identify TREM-2 in periodontal tissue, as well as the protein expression changes of TREM-1 and TREM-2 in periodontal tissues. MATERIAL AND METHODS: Gingival tissue sections were collected from 31 healthy subjects and 53 patients with chronic periodontitis. Immunohistochemistry and quantitative real-time polymerase chain reaction were employed to evaluate the protein and mRNA expression of these receptors in gingival tissues. The recorded clinical parameters were probing depth, clinical attachment loss, plaque index and bleeding on probing. RESULTS: In addition to myeloid cells in gingival connective tissues, TREM-1 and TREM-2 were also found expressed in gingival epithelial cells. In particular, TREM-1 was detected in almost all gingival epithelium from both healthy and inflamed biopsies. The expression levels of TREM-1 and TREM-2 were significantly increased in the periodontitis group compared to the healthy group. Increased levels of these receptors are to be positively correlated with site-specific periodontal parameters. CONCLUSION: The increased expression of TREM-1 and TREM-2 levels in periodontitis may confer diagnostic and potential therapeutic targets as well as indicating their association with the clinical severity of the disease.

Rhinoviruses and Their Receptors: Implications for Allergic Disease.

Human rhinoviruses (RVs) are picornaviruses that can cause a variety of illnesses including the common cold, lower respiratory tract illnesses such as bronchitis and pneumonia, and exacerbations of asthma. RVs are classified into three species, RV-A, B, and C, which include over 160 types. They utilize three major types of cellular membrane glycoproteins to gain entry into the host cell: intercellular adhesion molecule 1 (ICAM-1) (the majority of RV-A and all RV-B), low-density lipoprotein receptor (LDLR) family members (12 RV-A types), and cadherin-related family member 3 (CDHR3) (RV-C). CDHR3 is a member of cadherin superfamily of transmembrane proteins with yet unknown biological function, and there is relatively little information available about the mechanisms of RV-C interaction with CDHR3. A coding single nucleotide polymorphism (rs6967330) in CDHR3 could promote RV-C infections and illnesses in infancy, which could in turn adversely affect the developing lung to increase the risk of asthma. Further studies are needed to determine how RV infections contribute to pathogenesis of asthma and to develop the optimal treatment approach to control asthma exacerbations.

Identification of AcMNPV GP64-binding proteins through a combinational use of a self-biotinylated virus and the cross-linking method.

Baculoviruses are potential vectors of gene therapy for the ability to transfer gene high efficiently into mammalian cells. However, cell membrane proteins which interact with baculoviral glycoproteins have not been identified. In this study, we developed a self-biotinylated AcMNPV bearing biotinylated GP64 glycoproteins. This recombinant virus demonstrated the capability to infect insect cells and to transduct mammalian cells. Using this biotinylated virus, a protein >170Kda which could specifically interact with GP64 proteins was identified from virus transducted BHK-21 cells through cross-linking and streptavidin purification. Our study provides a useful approach for identifying cell membrane proteins that interact with baculovirus surface proteins or proteins involved in virus attachment.

Eruptive and functional changes in periodontal ligament fibroblast orientation in CD44 wild-type vs. knockout mice.

BACKGROUND AND OBJECTIVE: Periodontal ligament (PDL) fibroblasts establish principal fibers of the ligament during tooth eruption, and maintain these fibers during occlusion. PDL development and occlusal adaptation includes changes in the orientation of PDL fibroblasts; however, the mechanism for these changes in orientation is unclear. The objective of this study was to compare PDL fibroblast orientation in different stages corresponding with first molar eruption and occlusion in CD44 wild-type (WT) and knockout (KO) mice. MATERIAL AND METHODS: CD44 WT and KO mice were raised to six postnatal stages corresponding with first molar (M1 ) eruption (postnatal day 8, 11, 14 and 18) and occlusion (postnatal day 26 and 41). Coronal sections of the first mandibular molar (M1 ) were prepared and the orientation of fibroblasts in the cervical root region was measured. Angle measurements were compared across developmental stages and between strains using Watson-Williams F-test (oriana software) and ANCOVA. RESULTS: PDL fibroblast orientation increased significantly in CD44 WT (9-87°) and KO mice (14-93°; p ≤ 0.05) between intraosseous eruption (day 11), mucosal penetration (day 14) and preocclusal eruption (day 18); however, the PDL fibroblast orientation did not change significantly with the onset of occlusion (day 26) or continued function (day 41). Within each strain, the variance in fibroblast orientation during preocclusal eruption (day 18) was significantly higher than the variance of all other time points (p < 0.0005). CD44 WT and KO mice showed a similar pattern of PDL development and eruption with a significant difference in CD44 WT vs. KO fibroblast orientations only during early function (day 26, 92° vs 116°; p = 0.05). CONCLUSIONS: The development of PDL fibroblast orientation is highly similar between CD44 WT and KO mice. Between early (day 11) and late (day 18) eruptive stages PDL fibroblast orientation increases, corresponding with the upward movement of M1 . The PDL fibroblast orientation established in preocclusal eruption (day 18) is maintained during early (day 26) and late (day 41) stages of occlusal function, suggesting that PDL cells adapt to mechanical loads in the oral cavity before M1 occlusion.

Human cytokeratin 1 plays a role in the interaction of Pteropine orthoreovirus with Hek293 cells but not HeLa cells.

Pteropine orthoreovirus (PRV) causes respiratory tract infections in humans. Despite its emergence as a zoonotic and respiratory virus, little is known about its cell tropism, which hampers progress in fully understanding its pathogenesis in humans. Hek293 cells are most susceptible to PRV infection, while HeLa cells are the least. Human cytokeratin 1 (CK1) was identified as the protein that interacts with PRV. The immunofluorescence assay and qPCR results revealed prior treatment with anti-CK1 may provide Hek293 cells protection against PRV. The KRT1-knockout Hek293 cells were less susceptible to PRV infection. Further study into the pathogenesis of PRV in humans is needed.

Porphyromonas gingivalis, Treponema denticola and toll-like receptor 2 are associated with hypertensive disorders in placental tissue: a case-control study.

AIM(S): To explore the associations between the presence of periodontal pathogens and the expression of toll-like receptors (TLR-2 and TLR-4) in the placental tissue of patients with hypertensive disorders compared to the placentas of healthy normotensive patients. MATERIAL AND METHODS: A case-control study was performed. From a cohort composed of 126 pregnant women, 33 normotensive healthy pregnant women were randomly selected, and 25 cases of patients with hypertensive disorders of pregnancy, including gestational hypertension and pre-eclampsia, were selected. Placental biopsy was obtained after aseptic placental collection at the time of delivery. All of the samples were processed and analysed for the detection of Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Treponema denticola and Tannerella forsythia using the polymerase chain reaction (PCR) technique. Determination of the expressions of TLR-2 and TLR-4 was performed in samples of total purified protein isolated from placental tissues and analysed by ELISA. The data were assessed using descriptive statistics. The associations among variables were estimated through multiple logistic regression models and the Mann-Whitney test to evaluate the differences between the two groups. RESULTS: A significant increase was observed in the expression of TLR-2 in the placentas of patients with hypertensive disorders (p = 0.04). Additionally, the multiple logistic regression models demonstrated an association between the presence of T. denticola and P. gingivalis in placental tissues and hypertensive disorders (OR: 9.39, p = 0.001, CI 95% 2.39-36.88 and OR: 7.59, p = 0.019, CI 95% 1.39-41.51, respectively). CONCLUSIONS: In the present study, pregnant women with periodontal disease presented an association in the placental tissue between the presence of T. denticola and P. gingivalis and hypertensive disorders. Additionally, increased expression of TLR-2 was observed. However, further studies are required to determine the specific roles of periodontal pathogens and TLRs in the placental tissue of patients with pregnancy-related hypertensive disorders.

A New Cell Signal Regulation Theory: Blocking the Signal from being Reactivated Before a New Activation.

Currently, the concept of cell signal regulation has two basic steps: activation and deactivation. There are many activators of the signals in vivo (e.g., hormones, growth factors, signal cross-talk, etc.). According to the two step theory, cell signals are expected to keep activating, deactivating, activating, deactivating,……However, cell signals in vivo are usually quiescent for the most of the time. This indicates that, there is a mechanism that blocks the signal from being reactivated for a certain period of time after an activation is terminated. Here, we propose that cell's signal regulation consists of three basic steps: activation, deactivation, and blocking the signal from being reactivated for a certain period of time until cells are ready for a new activation. We explain the third step of regulation using GH/STAT5 and insulin/AKT signal pathways as examples, and discuss why the third step in regulation is important for cell homeostasis and the development of diseases.

Critical clinical gaps in cancer precision nanomedicine development.

Active targeting strategy is adopted in nanomedicine for cancer treatment. Personalizing the nanomedicine in accordance with patients' omics, under the precision medicine platform, is met with challenges in targeting ligand and matrix material selection at nanoformulation stage. The past 5-year literatures show that the nanoparticulate targeting ligand and matrix material are not selected based upon the cancer omics profiles of patients. The expression of cancer cellular target receptors and metabolizing enzymes is primarily influenced by age, gender, race/ethnic group and geographical origin of patients. The personalized perspective of a nanomedicine cannot be realised with premature digestion of matrix and targeting ligand by specific metabolizing enzymes that are overexpressed by the patients, and unmatched targeting ligand to the majority of cell surface receptors overexpressed in cancer. Omics analysis of individual metabolizing enzyme and cancer cell surface receptor expressed in cancer facilitates targeting ligand and matrix material selection in nanomedicine development.