3D Visualization of Dynamic Cellular Reaction of Pulpal CD11c+ Dendritic Cells against Pulpitis in Whole Murine Tooth

In dental pulp, diverse types of cells mediate the dental pulp immunity in a highly complex and dynamic manner. Yet, 3D spatiotemporal changes of various pulpal immune cells dynamically reacting against foreign pathogens during immune response have not been well characterized. It is partly due to the technical difficulty in detailed 3D comprehensive cellular-level observation of dental pulp in whole intact tooth beyond the conventional histological analysis using thin tooth slices. In this work, we validated the optical clearing technique based on modified Murray’s clear as a valuable tool for a comprehensive cellular-level analysis of dental pulp. Utilizing the optical clearing, we successfully achieved a 3D visualization of CD11c+ dendritic cells in the dentin-pulp complex of a whole intact murine tooth. Notably, a small population of unique CD11c+ dendritic cells extending long cytoplasmic processes into the dentinal tubule while located at the dentin-pulp interface like odontoblasts were clearly visualized. 3D visualization of whole murine tooth enabled a reliable observation of these rarely existing cells with a total number less than a couple of tens in one tooth. These CD11c+ dendritic cells with processes in the dentinal tubule were significantly increased in the dental pulpitis model induced by mechanical and chemical irritation. Additionally, the 3D visualization revealed a distinct spatial 3D arrangement of pulpal CD11c+ cells in the pulp into a front-line barrier-like formation in the pulp within 12 h after the irritation. Collectively, these observations demonstrated the unique capability of optical clearing-based comprehensive 3D cellular-level visualization of the whole tooth as an efficient method to analyze 3D spatiotemporal changes of various pulpal cells in normal and pathological conditions.

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Optical clearing based cellular-level 3D visualization of intact lymph node cortex

Biomedical Optics Express ◽

10.1364/boe.6.004154 ◽

2015 ◽

Vol 6 (10) ◽

pp. 4154 ◽

Cited By ~ 17

Author(s):

Eunjoo Song ◽

Howon Seo ◽

Kibaek Choe ◽

Yoonha Hwang ◽

Jinhyo Ahn ◽

...

Keyword(s):

Lymph Node ◽

3D Visualization ◽

Cellular Level ◽

Optical Clearing

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Co-increase of Nerve Fibers and HLA-DRand/or Factor-XIIIa-expressing Dendritic Cells in Dentinal Caries-affected Regions of the Human Dental Pulp: An Immunohistochemical Study

Journal of Dental Research ◽

10.1177/00220345990780100401 ◽

1999 ◽

Vol 78 (10) ◽

pp. 1596-1608 ◽

Cited By ~ 32

Author(s):

K. Sakurai ◽

T. Okiji ◽

H. Suda

Keyword(s):

Dendritic Cells ◽

Dental Pulp ◽

Coagulation Factor ◽

Nerve Fibers ◽

Factor Xiiia ◽

Morphological Evidence ◽

Immunoperoxidase Staining ◽

Dentinal Tubule ◽

Reparative Dentin ◽

Carious Teeth

Neuro-immune interaction has been suggested to play some modulatory role in the immunodefense of the dentin/pulp complex. In this study, we performed a simultaneous immunohistochemical observation of neural elements and pulpal dendritic cells (PDCs) on human carious teeth, to obtain morphological evidence for neuro-immune interaction in response to dentinal tubule-derived carious stimuli. Human third molars bearing a pulp-exposure-free caries lesion were studied. Immunoperoxidase staining was performed with anti-HLA-DR, anti-coagulation factor XIIIa, and anti-CD14 as PDC markers, and anti-low-affinity nerve growth factor receptor (NGFR), anti-protein gene products 9.5, and anti-calcitonin gene-related peptide as nerve markers. The carious teeth usually exhibited localized accumulation of both PDCs and nerve fibers immunoreactive to each marker, in the para-odontoblastic region corresponding to the pulpal end of carious dentinal tubules. Semi-quantitative digital densitometry revealed that pixel numbers corresponding to factor-XIIIa- and NGFRimmunoreactivity were significantly higher in the carious regions than those in the non-carious regions of the same teeth as well as those in the corresponding regions of intact teeth. Classification of specimens with respect to caries depth showed that the co-increase was most apparent in teeth with superficial caries. The increase of PDCs was less pronounced in carious teeth with reparative dentin. These findings suggest that both pulpal nerves and PDCs respond promptly and actively to dentinal tubule-derived carious stimuli. The synchronized accumulation of the two structures suggests an increased opportunity for neuro-immune interaction that may be of significance in the modulation of pathological processes in the dental pulp.

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Appearance and distribution of dendritic cells and macrophages in dental pulp during early postnatal morphogenesis of mouse mandibular first molars

Histochemistry and Cell Biology ◽

10.1007/s004180050407 ◽

1999 ◽

Vol 112 (3) ◽

pp. 193-204 ◽

Cited By ~ 11

Author(s):

Eichi Tsuruga ◽

Y. Sakakura ◽

Toshihiko Yajima ◽

Noriyuki Shide

Keyword(s):

Dendritic Cells ◽

Dental Pulp

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Study on application of optical clearing technique in skin diseases

Journal of Biomedical Optics ◽

10.1117/1.jbo.17.11.115003 ◽

2012 ◽

Vol 17 (11) ◽

pp. 115003 ◽

Cited By ~ 8

Author(s):

Hao Shan ◽

Yanmei Liang ◽

Jingyi Wang ◽

Yan Li

Keyword(s):

Skin Diseases ◽

Optical Clearing ◽

Clearing Technique

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Cancer Stem Cells and Epithelial Ovarian Cancer

Journal of Oncology ◽

10.1155/2010/105269 ◽

2010 ◽

Vol 2010 ◽

pp. 1-9 ◽

Cited By ~ 17

Author(s):

Sheetal Dyall ◽

Simon A. Gayther ◽

Dimitra Dafou

Keyword(s):

Ovarian Cancer ◽

Stem Cells ◽

Stem Cell ◽

Cancer Stem Cells ◽

Cancer Stem Cell ◽

Epithelial Ovarian Cancer ◽

Primary Tumour ◽

Cellular Level ◽

Small Population ◽

Novel Target

The cancer stem cell hypothesis is becoming more widely accepted as a model for carcinogenesis. Tumours are heterogeneous both at the molecular and cellular level, containing a small population of cells that possess highly tumourigenic “stem-cell” properties. Cancer stem cells (CSCs), or tumour-initiating cells, have the ability to self-renew, generate xenografts reminiscent of the primary tumour that they were derived from, and are chemoresistant. The characterisation of the CSC population within a tumour that drives its growth could provide novel target therapeutics against these cells specifically, eradicating the cancer completely. There have been several reports describing the isolation of putative cancer stem cell populations in several cancers; however, no defined set of markers has been identified that conclusively characterises “stem-like” cancer cells. This paper highlights the current experimental approaches that have been used in the field and discusses their limitations, with specific emphasis on the identification and characterisation of the CSC population in epithelial ovarian cancer.

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Turnover of Dermal Dendritic Cells.

Blood ◽

10.1182/blood.v106.11.2283.2283 ◽

2005 ◽

Vol 106 (11) ◽

pp. 2283-2283

Author(s):

Milena Bogunovic ◽

Florent Ginhoux ◽

Martine Loubeau ◽

Miriam Merad

Keyword(s):

Dendritic Cells ◽

Steady State ◽

Chemokine Receptors ◽

Uv Light ◽

Small Population ◽

Specific Marker ◽

Chimeric Mice ◽

Complete Replacement ◽

Surface Marker Expression ◽

High Level

Abstract We have previously shown that in steady state conditions, epidermal Langerhans cells (LCs) derive from a radio-resistant precursor that self-renew in the skin throughout life (Merad et al. 2002). In the current study we addressed the mechanisms that regulate the turnover of other subsets of skin antigen presenting cells (APCs). Based on cell surface marker expression, we characterized two populations of APCs in the mice dermis including CD11c+ CD11b+ cells and CD11b+ CD11c- cells (likely corresponding to dermal dendritic cells (DDCs) and macrophages respectively). To explore the turnover of DDCs we reconstituted lethally irradiated CD45.2+ C57BL/6 mice with bone marrow (BM) cells isolated from CD45.1+ C57BL/6 congenic mice and followed the replacement of host DDCs by donor DDCs. Interestingly, we found that despite 95% blood and BM chimerism, 20% of CD11c+ DDCs remained of host (CD45.2+) origin for at least 1 year after transplant, while 80% of DDCs and dermal macrophages appeared to be of donor (CD45.1+) origin as early as at 4 weeks after transplant. The radio-resistant CD11c+ cells homogenously expressed CD11b, F4/80, high-level MHC II and the co-stimulatory molecules CD40 and CD86, while only 10% of them expressed the LC specific marker (langerin) suggesting that these cells represent migrating LCs. To explore whether radio-resistant DDCs are able to proliferate locally, we administered bromodeoxyuridine (BrdU) to (CD45.1+ BM > CD45.2+ recipient) chimeric mice, 8 weeks after transplant. Because at the time of BrdU administration, blood cells in chimeric mice were of donor origin, incorporation of BrdU in host DDCs should reflect local proliferation. Indeed, we found that 30% of host DDCs incorporate BrdU after two weeks of BrdU labeling suggesting that radio-resistant DDCs proliferate in situ. In contrast to steady state conditions, exposure of chimeric mice to ultraviolet (UV) light led to complete replacement of the remaining host DDCs by circulating donor DDCs. To explore the nature of the chemokines playing a role in the recruitment of circulating DDCs to inflamed skin, we reconstituted lethally irradiated CD45.1+ mice with a 1:1 mixture of CD45.1+ autologous BM cells and CD45.2+ BM cells deficient in the chemokine receptors CCR2 or CCR6. Two weeks after transplant, we exposed the mice to UV light and followed the recruitment of mutant and wild type DDCs in the skin. Importantly, we found that 3 weeks after UV exposure, the % of CD45.2+ CCR2−/− DDCs was 8 fold lower than CD45.1+ CCR2+/+ DDCs, while there was no difference between the % of CCR6−/− and CCR6+/+ DDCs. These results suggest that the recruitment of donor DDCs is dependent on the chemokine receptor CCR2 but not CCR6, while CCR2 and CCR6 were required for the recruitment of circulating LC precursors. Taken together, our results suggest that, similar to LCs, a small population of DDCs is radio-resistant and proliferate in quiescent skin. Whether this subset of DDCs represents a local precursor for epidermal LCs will be the subject of further studies.

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Residual Lymphocytes in GM-CSF and IL-15 Differentiated Monocyte-Derived Dendritic Cells Enables Cytotoxic T Lymphocyte Responses.

Blood ◽

10.1182/blood.v110.11.4907.4907 ◽

2007 ◽

Vol 110 (11) ◽

pp. 4907-4907

Author(s):

Melinda Y. Hardy ◽

Andrew J. Kassianos ◽

Ray Wilkinson ◽

Annelie Vulink ◽

Derek N.J. Hart ◽

...

Keyword(s):

Dendritic Cells ◽

T Lymphocyte ◽

Cytotoxic T Lymphocyte ◽

Small Population ◽

Lymphoid Cells ◽

Gm Csf ◽

Hla Dr ◽

Ifn Γ ◽

Human Dendritic Cells ◽

Ctl Responses

Abstract We investigated the capacity of IL-15 to differentiate human dendritic cells (DC) from monocytes in the presence of GM-CSF (IL-15 MoDC) and compared them with MoDC differentiated in IL-4 and GM-CSF (IL-4 MoDC) as used in many immunotherapy protocols. IL-15 MoDC expressed higher levels of CD40 and HLA-DR and importantly, induced MART-1 specific cytotoxic T lymphocyte (CTL) responses with superior lytic capacity, when compared to IL-4 MoDC. In response to activation, IL-15 MoDC secreted high levels of IFN-γbut low or no IL-12, whereas IL-4 MoDC secreted high IL-12 but low or no IFN-γ. Using an IFN-γ blocking antibody, we demonstrated that IFN-γ production by the IL-15 MoDC did not account for the superior CTL responses induced. Despite immunoselecting monocytes to greater than 97% purity prior to DC differentiation, we noticed a small population (1–2%) of CD56+ and CD3+ lymphocytes in the IL-15 MoDC preparations that were less prominent in IL-4 MoDC differentiated from the same monocytes. Removal of the residual lymphocytes from monocytes prior to differentiation into IL-15 MoDC diminished their capacity to induce CTL but did not affect the expression of HLA-DR or CD40. These data suggest that IL-15-dependent cross-talk between the small lymphoid populations present and DC, during DC differentiation from monocytes results in superior CTL priming that is independent of IL-12 and IFN-γ. Based on these results, appropriately manufactured IL-15 MoDC preparations containing defined numbers of lymphoid cells should be considered for immunotherapy.

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Breathing with Phox2b

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2009.0085 ◽

2009 ◽

Vol 364 (1529) ◽

pp. 2477-2483 ◽

Cited By ~ 28

Author(s):

Véronique Dubreuil ◽

Jacques Barhanin ◽

Christo Goridis ◽

Jean-François Brunet

Keyword(s):

Reverse Genetics ◽

Human Genetics ◽

Respiratory Rhythm ◽

Neuronal Population ◽

Cellular Level ◽

Small Population ◽

Respiratory Neurons ◽

Retrotrapezoid Nucleus ◽

Parafacial Respiratory Group

In the last few years, elucidation of the architecture of breathing control centres has reached the cellular level. This has been facilitated by increasing knowledge of the molecular signatures of various classes of hindbrain neurons. Here, we review the advances achieved by studying the homeodomain factor Phox2b , a transcriptional determinant of neuronal identity in the central and peripheral nervous systems. Evidence from human genetics, neurophysiology and mouse reverse genetics converges to implicate a small population of Phox2b -dependent neurons, located in the retrotrapezoid nucleus, in the detection of CO 2 , which is a paramount source of the ‘drive to breathe’. Moreover, the same and other studies suggest that an overlapping or identical neuronal population, the parafacial respiratory group, might contribute to the respiratory rhythm at least in some circumstances, such as for the initiation of breathing following birth. Together with the previously established Phox2b dependency of other respiratory neurons (which we review briefly here), our new data highlight a key role of this transcription factor in setting up the circuits for breathing automaticity.

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