Apolipoprotein E controls Dectin-1-dependent development of monocyte-derived alveolar macrophages upon pulmonary ß-glucan-induced inflammatory adaptation.

The lung is constantly exposed to the outside world and optimal adaptation of immune responses is crucial for efficient pathogen clearance. However, mechanisms that lead to lung-associated macrophages' functional and developmental adaptation remain elusive. To reveal such mechanisms, we developed a reductionist model of environmental intranasal ß-glucan exposure, allowing for the detailed interrogation of molecular mechanisms of pulmonary macrophage adaptation. Employing single-cell transcriptomics, high-dimensional imaging and flow cytometric characterization paired with in vivo and ex vivo challenge models, we reveal that pulmonary low-grade inflammation results in the development of apolipoprotein E (ApoE)-dependent monocyte-derived alveolar macrophages (ApoE+CD11b+ AMs). ApoE+CD11b+ AMs expressed high levels of CD11b, ApoE, Gpnmb and Ccl6, were glycolytic, highly phagocytic and produced large amounts of interleukin-6 upon restimulation. Functional differences were cell intrinsic, and myeloid cell-specific ApoE ablation inhibited Ly6c+ monocyte to ApoE+CD11b+ AM differentiation dependent on macrophage colony-stimulating factor secretion, promoting ApoE+CD11b+ AM cell death and thus impeding ApoE+CD11b+ AM maintenance. In vivo, ß-glucan-elicited ApoE+CD11b+ AMs limited the bacterial burden of Legionella pneumophilia after infection and improved the disease outcome in vivo and ex vivo in a murine lung fibrosis model. Collectively these data identify ApoE+CD11b+ AMs generated upon environmental cues, under the control of ApoE signaling, as an essential determinant for lung adaptation enhancing tissue resilience.

Microvascular regulatory role and increased expression of vascular endothelial growth factor receptor type 2 in experimental gingivitis.

OBJECTIVE: The aim of the present study was to investigate the possible microvascular regulatory role of vascular endothelial growth factor receptor type 2 (VEGFR2) in experimental gingivitis in rats. BACKGROUND: Our previous results demonstrated that functionally active VEGFR2s are located in the venules of rat gingiva. While there is no remarkable endogenous gingival VEGF production under normal circumstances, exogenous VEGF, via VEGFR2, shows venodilatory effects. We assumed that VEGF plays an important role in vasoregulatory processes (vasodilation, increased permeability, angiogenesis) of gingival inflammation. METHODS: Gingivitis was induced by placing ligatures and composite material around and between the lower incisors of anesthetized Wistar rats next to the gingival margin. Seven days later, VEGFR2 antagonist (ZM323881), was dripped upon the labial gingiva next to the lower incisors. Diameter changes of the selected gingival venules were measured by vital microscopy. Animals with healthy gingiva served as controls. Venule diameter changes were compared to the baseline and to control groups (no ligature). Immunohistochemical and Western blot analysis for VEGFR2 were utilized. RESULTS: After 15, 30 and 60 min of local application of ZM323881, there was a significant venoconstriction in the inflamed gingiva compared to the baseline, while no change was recorded in controls. Endothelium, smooth muscle cells and pericytes of the gingivitis group showed increased VEGFR2 expression. CONCLUSION: Our findings suggest that there is an increased VEGF production in gingivitis, which may play an important role in vasodilation of rat gingival venules.

[Blood flow in marginal gingiva as measured with laser Doppler flowmetry]. / A marginális gingiva véráramlásának vizsgálata lézer Doppler-áramlásméróvel.

The clinical application of laser Doppler flowmetry (LDF) provides data on the blood flow of marginal gingiva (GBF) at different dental regions. The aim of the present investigation was to study the GBF at different teeth. Ten adult subjects with good oral hygiene were involved in this experiment. In a series of experiments GBF was recorded bilaterally positioned the flow probe on three sites 1 mm above the marginal gingiva of the upper and lower central incisors. In another series of investigations the same technique was used for GBF registrations at six permanent teeth on the right side (11-16; according to FDI notation). The mean blood flow values obtained at 3 different places of marginal gingiva of the front teeth were nearly identical, therefore the data were pooled. The GBF values registered at the six upper right teeth also showed close similarity (p > 0.30). No differences were observed between the mean GBF values at the left and right central incisors (p > 0.80). There was a significant gingival blood flow value elevation at lower incisors as compared to that in the upper jaw (p < 0.05). Our results indicate that there is homogeneous marginal blood perfusion both in the maxillary and mandibula healthy gingiva. However, difference exists in blood supply between the upper and lower jaws in favor of mandibular marginal gingiva.

[Blood flow measurements in human oral tissues with laser Doppler flowmetry]. / A szájüregi képletek keringésének vizsgálata lézer Doppler-áramlásméróvel humán egyedekben.

In our examination the non-invasive laser Doppler flowmetry (LDF) was used to determine the blood flow in the tooth pulp (PBF) and in the gingiva (GBF) in humans. Perfusion values were collected by the means of LDF probe adjusted manually or by manipulator assisted fixation. Significantly lower GBF rate was detected by hand-held probe as compared to that obtained by manipulator assisted fixation (p < 0.001). No significant difference was observed for the PBF rates (p > 0.9). The coefficient of variation determined for GBF was higher (0.32 +/- 0.09) in case of manual procedure than at manipulator assisted (0.13 +/- 0.02) fixation mode. In contrary, the value of variation coefficient of PBF data obtained by hand-held probe was lower as compared to that calculated from data of manipulator fixation (0.16 +/- 0.08 v. 0.22 +/- 0.1; p < 0.05). These data indicate that while the manual fixation for measuring PBF seems to be satisfactory, for the accurate determination of GBF rate application of manipulator is essential.

Protective effects of mercaptoethylguanidine, a selective inhibitor of inducible nitric oxide synthase, in ligature-induced periodontitis in the rat.

1. Excessive production of nitric oxide (NO), and the generation of peroxynitrite have been implicated in various proinflammatory conditions. In the present study, using mercaptoethylguanidine (MEG), a selective inhibitor of iNOS and a peroxynitrite scavenger, we investigated the role of iNOS and peroxynitrite in a rat model of periodontitis. 2. Periodontitis was produced in rat by a ligature of 2/0 braided silk placed around the cervix of the lower left 1st molar. Animals were then divided into two groups: one group of rats was treated with MEG (30 mg kg(-1), i.p., 4 times per day for 8 days), animals in the other group received vehicle. At day 8, the gingivomucosal tissue encircling the mandibular 1st molars was removed on both sides from ligated and sham operated animals for inducible nitric oxide synthase (iNOS) activity assay and for immunocytochemistry with anti-iNOS serum. Plasma extravasation was measured with the Evans blue technique. Alveolar bone loss was measured with a videomicroscopy. 3. Ligation caused a significant, more than 3 fold increase in the gingival iNOS activity, whereas it did not affect iNOS activity on the contralateral side, when compared to sham-operated animals. Immunohistochemical analysis revealed iNOS-positive macrophages, lymphocytes and PMNs in the connective tissue and immunoreactive layers of epithelium on side of the ligature, and only a few iNOS reactive connective tissue cells on the contralateral side [corrected]. Ligation significantly increased Evans blue extravasation in gingivomucosal tissue and alveolar bone destruction compared to the contralateral side. MEG treatment significantly reduced the plasma extravasation and bone destruction. 4. The present results demonstrated that ligature-induced periodontitis increases local NO production and that MEG treatment protects against the associated extravasation and bone destruction. Based on the present data, we propose that enhanced formation of NO and peroxynitrite plays a significant role in the pathogenesis of periodontitis.

Effects of streptozotocin-induced diabetes on neurogenic inflammation of gingivomucosal tissue in rat.

It has been suggested that the unmyelinated small diameter afferent nociceptive C-fibres are impaired in diabetes mellitus. We have recently demonstrated that these fibres are the prerequisite for neurogenic inflammation induced by mechanical or chemical irritations. These experiments were designed to characterize the neurogenic inflammatory responses of gingivomucosal tissue in the early phase of experimental induced diabetes mellitus in rat. Effect of dental ligature on the gingivomucosal (GM) vascular permeability was studied in control rats and in rats pretreated with streptozotocin at d 7 and 14 following streptozotocin administration. In separate groups of control and streptozotocin diabetic rats studies were also performed to investigate the effect of local capsaicin application on GM vascular permeability on d 14. Vascular permeability was assessed by means of Evans blue extravasation. The ligature placed around the mandibular left first molar caused a significant increase vascular permeability of GM tissue on the ipsilateral side on both d 7 and 14 after the ligation in control rats. In streptozotocin diabetic rats on d 7, there was also a significant elevation of Evans blue extravasation in the tissue tested on the ligature side. However, on d 14 the ligation failed to produce any changes in Evans blue extravasation on the ipsilateral side, i.e. no difference in GM vascular permeability could be recorded between the two sides in streptozotocin diabetic rats. Topical capsaicin administration produced significant Evans blue extravasation in GM tissue of control rats compared to that observed in diabetic rats on d 14 after streptozotocin treatment. Electron microscopic and light microscopic studies demonstrated fibre degeneration of the C neurones and less inflammatory cells in streptozotocin-induced diabetes in the gingivomucosal tissue. These findings appear to indicate that the inflammatory responses induced by mechanical (dental ligature) and/or chemical irritants (topical application of capsaicin) in the gingivomucosal tissue are altered in streptozotocin diabetic rats and this alteration is due to the diabetes-induced damage to the unmyelinated C fibres.

Effect of substance P administration on vascular permeability in the rat oral mucosa and sublingual gland.

Neuropeptides, including substance P (SP) may play a role in neurogenic inflammation. Although SP-immunoreactive (SP-IR) axons are known to be present within the oral mucosa (OM) and salivary glands, the functional significance of SP in oral mucosa and sublingual salivary gland (SLG) is not fully understood. The present experiments were carried out to study the effects of SP infused into the left common carotid artery on vascular permeability in the OM and in the SLG of male rats. Vascular permeability was assessed on the basis of Evans Blue extravasation. Separate groups of animals received histamine (H1) receptor antagonist (chloropyramine, 10 mg kg-1 i.v.) or prostaglandin synthesis inhibitor (indomethacin, 4 mg kg-1 i.v.) prior to SP infusions. Infusion of SP in doses of 30 and 74 pmol min-1 increased the vascular permeability of OM by 162.3 +/- 16.3% (n = 8, p < 0.05) and 482.7 +/- 46.7% (n = 8, p < 0.001), respectively. SP in a dose of 15 pmol min-1 did not increase Evans Blue extravasation in OM (38.3 +/- 4.0 micrograms g-1, n = 8, compared to the control: 44.0 +/- 7.9 micrograms g-1, n = 8, NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Neurogenic component in ligature-induced periodontitis in the rat.

Effect of ligation on the vascular permeability in the gingiva and alveolar mucosa encircling the mandibular left 1st molar was studied in rats with and without capsaicin pretreatment. Vascular permeability was assessed by the Evans blue extravasation. Ligation caused a significant augmentation in vascular permeability of the gingivomucosal tissue at day 8 (right: 18.14 +/- 1.68 micrograms g-1; left (ligature): 38.21 +/- 2.43 micrograms g-1, n = 8, p < 0.001) and at day 14 (right: 20.31 +/- 1.71 micrograms g-1: left (ligature): 36.98 +/- 2.73 micrograms g-1, n = 8, p < 0.001). 4 days after ligation, no difference could be observed in vascular permeability in the oral mucosa of the ligated side (left: 23.14 +/- 1.21 micrograms g-1) as compared to the side without ligature (right: 23.5 +/- 1.45 micrograms g-1, n = 8, NS). There was no elevation of vascular permeability of gingivomuscosal tissue around the ligation in rats pretreated with capsaicin either in newborn age (right: 23.92 +/- 1.76 micrograms g-1; left (ligature): 23.51 +/- 2.16 micrograms g-1, n = 8, NS) or in adult age (right: 20.61 +/- 1.62 micrograms g-1; left (ligature): 20.85 +/- 1.07 micrograms g-1, n = 8, NS). Light microscopical studies of oral mucosa revealed, that 8 and 14 days after the ligature placed around the mandibular left 1st molar of the rat, there resulted an accumulation of inflammatory cells in the connective tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of substance P (SP) in development of symptoms of neurogenic inflammation in the oral mucosa of the rat.

In the present series of investigations we first studied the local effects of exogenous substance P (SP) on the hallmarks of neurogenic inflammation, i.e. vascular permeability and blood flow, in the oral mucosa of the rat. Pretreatment with capsaicin was shown to attenuate the symptoms of neurogenic inflammation; therefore, the distribution of nerve fibers displaying substance P-like immunoreactivity (SP-IR) in the mandibular mucosa was also assessed in control rats and in animals pretreated with capsaicin both neonatally and in adulthood using immunohistochemical techniques. The application of SP at a dose of 7.5 nmol resulted in an almost 70% increase of vascular permeability (NS) and the administration of a four-fold higher dose (30 nmol) produced about 150% increase in Evans blue extravasation compared with control values (p < 0.05). A similar increase (ca 146%) in vascular permeability was observed in response to 45 nmol SP solution (p < 0.05). While the 7.5 nmol SP-solution failed to affect blood flow, the 30 nmol SP significantly increased it by ca. 38% (p < 0.05). The administration of the 45 nmol SP solution resulted in a similar enhancement of blood flow (43%, p < 0.05). Capsaicin pretreatment performed either neonatally or in adulthood has reduced the number of SP-immunoreactive fibers in the oral mucosa. Our functional results suggest that SP may have a role in the experimentally-induced neurogenic inflammation of the oral mucosa in the rat. This is also supported by our finding that capsaicin pretreatment, known to decrease the number of SP-immunoreactive fibers in these tissues, reduced the symptoms of neurogenic inflammation.

Neurogenic inflammation and the oral mucosa.

Since the rôle of neurogenic inflammation in various pathological conditions is well-established, we presume that it also has great importance in the development of several inflammatory processes in the oral mucosa. In the course of experimentally-induced neurogenic inflammation of the oral mucosa in rats, vasodilation and plasma extravasation occur in the area supplied by unmyelinated capsaicin-sensitive fibres. Upon activating these fibres in the oral mucosa, mediators such as histamine, substance P (SP) and calcitonin gene related peptide (CGRP) are released from the peripheral terminals of afferent nerves causing characteristic symptoms of neurogenic inflammation. By histochemical techniques, SP- and CGRP-immunoreactive fibres have occurred in the area of the rat oral mucosa stimulated mostly in the free and attached gingiva around the molar teeth in the lower jaw. Capsaicin pretreatment performed neonatally or at adult age causes a loss of SP-immunoreactive fibres and prevents the vasodilatory responses, as well as the increase in vascular permeability elicited by the antidromic stimulation of the inferior alveolar nerve. As the transection of inferior alveolar nerve decreased the extravasation of Evans blue on the ipsilateral side and did not affect the capsaicin-induced enhancement in blood flow, we suggest that the two symptoms of the inflammation, i.e., the increased vascular permeability and decreased vascular resistance, should be produced by different mechanisms. The results of all the morphological and functional studies seem to confirm the possibility that there is an important neurogenic component of the inflammatory alterations caused by different mechanical and chemical stimuli in the oral mucosa.

Effect of substance P administration on vascular permeability in the rat dental pulp and submandibular gland.

The effects of substance P (SP) administration on vascular permeability were studied in the dental pulp (DP) of upper and lower incisors and in the submandibular gland (SMG) of male rats. Vascular permeability was assessed by means of extravasation of Evans blue dye. SP was diluted in 0.5% bovine serum albumin (BSE) and infused into the left common carotid artery. Separate groups of animals receive chloropyramine, an H1 histamine receptor antagonist (10 mg kg-1 i.v.) or indomethacin, a prostaglandin synthesis inhibitor (4 mg kg-1 i.v.) prior to SP infusions. Infusion of SP for 5 min increased plasma extravasation both in DP and SMG, with a threshold of about 30 pmol min-1 and 74 pmol min-1, respectively. Enhanced salivary secretion was also observed. Although the administration of 74 pmol min-1 of SP significantly lowered the systemic blood pressure, experimental hypotension elicited by haemorrhage did not influence vascular permeability in either organ tested. After chloropyramine administration the SP effect on vascular permeability in both DP and SMG was abolished. Indomethacin pretreatment failed to prevent the permeability-enhancing action of SP. Our results suggest that substance P increases both pulpal and glandular plasma extravasation in the rat indirectly, via the release of histamine and the activation of H1 histamine receptors.

[The role of histamine in neurogenic inflammations of the mouth mucosa in rats]. / A hisztamin szerepe a patkány-szájnyálkahártya neurogén inflammációjában.

The effect mechanism of neurogen inflammation incited by local capsaicin irritation was examined by the authors in the oral soft tissue of rats subsequent to antihistamin pretreatment. Furthermore, with the aid of muscarin blocking receptor also the parasympathetic component of the vasodilatation was examined. On basis of their results it was established that the vein-wall-permeability and microcircular changes observed in the course of the neurogen inflammation of the oral soft tissue partly expanded by the histamin through the H1-receptors. The H2-receptors as well as the parasympathetic fibers do not become activated in the process.

Effect of denervation on the neurogenic inflammation of the rat mandibular mucosa.

Effects of local exposure to capsaicin on the vascular permeability and blood flow were studied in the rat oral mucosa at days 2 and 14 after the unilateral transection of the inferior alveolar nerve (IAN). The distribution of nerve fibers displaying substance P (SP) and calcitonin gene-related peptide (CGRP) immunoreactivity (IR) in the mandibular mucosa was also assessed. While the capsaicin-induced augmentation in vascular permeability was about 50% (P less than 0.05) higher on the intact side (at both days 2 and 14) than on the denervated side, no difference in blood flow elevation was seen between the two sides. Transection of IAN caused only a slight reduction in the density of SP- and CGRP-IR fibers in the mucosa. It is concluded that in addition to the IAN fibers the mandibular mucosa examined also seems to be supplied by other sensory fibers. The presence of accessory trigeminal branches was also supported by immunohistochemical studies.

Role of histamine in the development of neurogenic inflammation of rat oral mucosa.

The mechanism of development of neurogenic inflammatory reaction induced by the topical application of capsaicin was studied in the oral mucosa of rats with or without histamine antagonist pretreatment. The existence of a cholinergic component of the vasodilation was investigated using a muscarinic receptor antagonist. Results indicated that the neurogenic inflammatory increases in vascular permeability and blood flow are mediated in part by H1-receptors, H2-receptors and cholinergic pathways are apparently not activated in these processes.

[The effect of the transsection of the inferior alveolar nerve on neurogenic inflammation of the mouth mucosa. II. Immunohistochemical studies]. / A nervus alveolaris inferior átmetszésének hatása a szájnyálkahártya neurogén gyulladása. II. Immunhisztokémiai vizsgálatok.

The distribution of nerve fibers displaying calcitonin gene-related peptide (CGRP) immunoreactivity in the mandibular mucosa was studied in control rats, in rats after unilateral transection of the inferior alveolar nerve (IAN), and in animals following cervical sympathetic denervation. According to the results, the transection of the peripheral branch of the IAN has no effect on the distribution of the CGRP immunoreactive nerves.

[The effect of transsection of the inferior alveolar nerve on neurogenic inflammation of the oral mucosa. I. Functional studies]. / A nervus alveolaris inferior átmetszésének hatása a szájnyálkahártya neurogén gyulladására. I. Funkcionális vizsgálatok.

Vascular effects of local capsaicin treatment has been studied on the 2nd and 14th days subsequent to unilateral inferior alveolar nerve (IAN) transection in the oral mucosa of rats. The results suggest, that the symptoms of neurogenic inflammation were influenced by the transection of IAN in a different way. The reason of that is the different mechanism in the development of the symptoms and/or the different compensatory capacity of the accessorial nerves.

Experimentally-induced neurogenic inflammation in the rat oral mucosa.

Blood flow and vascular permeability in the gingiva and alveolar mucosa of the lower jaw were studied after the antidromic (electric) or orthodromic (topical administration of capsaicin onto the oral mucosa) stimulation of the inferior alveolar nerve (IAN) in rats with or without capsaicin pretreatment. Blood flow was determined using 86Rb isotope dilution technique, while vascular permeability was assessed by the Evans blue extravasation. Both antidromic or orthodromic stimulation of the IAN increased blood flow and Evans blue extravasation. Capsaicin pretreatment abolished the increase in gingivo-mucosal blood flow and vascular permeability induced by nerve stimulation. The results of the present study appear to confirm the possibility of the development of neurogenic inflammation in the oral cavity. The mechanism observed may play a role in the development of certain inflammatory reactions in the oral mucosa frequently encountered in human clinical practice.