Analysis of etiology and clinical features of spontaneous downbeat nystagmus: a retrospective study

ObjectiveTo investigate the topical diagnosis, possible etiology and mechanism of spontaneous downbeat nystagmus (sDBN) patients with dizziness/vertigo.MethodsThe clinical features of dizziness/vertigo patients accompanied with DBN were retrospectively reviewed in the Vertigo Center of our hospital from January 2018 to March 2021. The clinical features of dizziness/vertigo patients accompanied with DBN were reviewed. Comprehensive VNG, bithermal caloric testing, video-head-impulse test (vHIT), vestibular-evoked myogenic potentials (VEMP), head magnetic resonance imaging (MRI), three-dimensional fluid-attenuated incersion recovery magnetic resonance imaging (3D-FLAIR MRI) in the inner ear, serum immunology and other examinations were to determine the lesion site, and analyze its possible etiology and mechanism.ResultsA total of 54 patients were included. Among them, 70.4% (n = 38) of DBN patients were diagnosed with episodic vestibular syndrome (EVS), 22.2% (n = 12) with chronic vestibular syndrome (CVS), and 7.4% (n = 4) with acute vestibular syndrome (AVS). Among all the patients, 51.9% of DBN patients had clear etiology, with central lesions of 29.6% and peripheral diseases of 22.2%. The most common diseases in DBN patients were cerebellar lesions (13.0%, n = 7) and vestibular migraine (13.0%, n = 7), followed by benign positional paroxysmal vertigo (7.4%, n = 4) and drug-related dizziness/vertigo (5.6%, n = 3). The other 48.1% of the patients had unknown etiology. 53.8% (14/26) of patients with idiopathic DBN had decreased semicircular canal function, with 42.9% (6/14) decreased posterior semicircular canal function. The posterior semicircular canal gain in DBN patients decreased compared to the anterior semicircular canal in the same conjugate plane. Patients with peripheral DBN were more prone to horizontal/torsional nystagmus during positional testing.ConclusionIn our study, DBN patients have a relative decrease in posterior semicircular canal gain, which is possibly a particular result found in a subset of downbeat nystagmus patients. The changes in nystagmus during positional testing may be helpful in distinguishing between peripheral and central causes

IL-17A Synergizes with IFN-γ to Upregulate iNOS and NO Production and Inhibit Chlamydial Growth

IFN-γ-mediated inducible nitric oxide synthase (iNOS) expression is critical for controlling chlamydial infection through microbicidal nitric oxide (NO) production. Interleukin-17A (IL-17A), as a new proinflammatory cytokine, has been shown to play a protective role in host defense against Chlamydia muridarum (Cm) infection. To define the related mechanism, we investigated, in the present study, the effect of IL-17A on IFN-γ induced iNOS expression and NO production during Cm infection in vitro and in vivo. Our data showed that IL-17A significantly enhanced IFN-γ-induced iNOS expression and NO production and inhibited Cm growth in Cm-infected murine lung epithelial (TC-1) cells. The synergistic effect of IL-17A and IFN-γ on Chlamydia clearance from TC-1 cells correlated with iNOS induction. Since one of the main antimicrobial mechanisms of activated macrophages is the release of NO, we also examined the inhibitory effect of IL-17A and IFN-γ on Cm growth in peritoneal macrophages. IL-17A (10 ng/ml) synergizes with IFN-γ (200 U/ml) in macrophages to inhibit Cm growth. This effect was largely reversed by aminoguanidine (AG), an iNOS inhibitor. Finally, neutralization of IL-17A in Cm infected mice resulted in reduced iNOS expression in the lung and higher Cm growth. Taken together, the results indicate that IL-17A and IFN-γ play a synergistic role in inhibiting chlamydial lung infection, at least partially through enhancing iNOS expression and NO production in epithelial cells and macrophages

A Pathogenic Role for FcγRI in the Immune Response against Chlamydial Respiratory Infection

FcγRI is an important cell surface receptor reported to be involved in multiple immune responses, although it has not yet been extensively studied in intracellular bacterial infections. Here, using a mouse model of C. muridarum respiratory infection, we were able to determine how FcγRI regulates the host resistance against chlamydial invasion. According to our findings, the chlamydial loads and pulmonary pathology were both reduced in FcγRI deficient (Fcgr1−/−) animals. Being infected, monocytes, macrophages, neutrophils, DCs, CD4+/CD8+ T cells, and effector Th1 subsets displayed increased FcγRI expression patterns. Altered infiltration of these cells in the lungs of Fcgr1−/− mice further demonstrated the regulation of FcγRI in the immune system and identified Th1 cells and macrophages as its target cell populations. As expected, we observed that the Th1 response was augmented in Fcgr1−/− mice, while the pro-inflammatory M1 macrophage polarization was constrained. These findings might indicate FcγRI as a potential regulator for host immunity and inflammatory response during chlamydial infection

The comparison of iNOS expression between IL-17A-neutralized mice and IgG2a-treated control mice.

<p>Mice were intranasally infected with 1×10³ IFUs of Cm. For neutralization of airway IL-17A, mice were intranasally administered with 10 µg of anti-mouse IL-17A mAb in 40 µl of PBS at 2 h post-infection (p.i.), followed by subsequent administration of the same dose of Ab at 48-h intervals. Infected control mice were administered intranasally with anti-murine IL-17A Ab isotype control (IgG2a) in the same schedule as anti-IL-17A delivery. Mice were monitored for body weight changes (A) and killed at day 7 after infection. The lungs were collected and analyzed for in vivo chlamydial growth (B) and gene expression (C and D) as described in <i>Materials and Methods</i>. D, densitometric analysis of iNOS mRNA RT-PCR products from lung in mice at day 7 postinfection. Values were normalized to β-actin and expressed as the mean ± SD with n = 5 for each point. **, p<0.01.</p

Effect of IL-17A and IFN-γ on iNOS gene expression, iNOS activity and NO production in RAW 264.7 cells.

<p>RAW 264.7 cells were cultured and treated with rmIL-17A (10 ng/ml) and/or rmIFN-γ (20 ng/ml) for 6 h. Cells were harvested, RNA was isolated and the mRNA level of iNOS was measured by RT-PCR. β-actin was used as an internal control. The representative gels were presented. The values of iNOS expression were normalized to β-actin and expressed as the mean ± SD of triplicate measurements (A). iNOS activity of cell lysates was detected by chromatometry method (B). NO concentration in cell culture supernatant was detected using NO nitrate reductase method (C). *P<0.05, **P<0.01, ***P<0.001.</p

Effect of IL-17A and IFN-γ on iNOS gene expression, iNOS activity and NO production in TC-1 cells.

<p>TC-1 cells were cultured and treated with rmIL-17A (10 ng/ml) and/or rmIFN-γ (20 ng/ml) for 6 h. Cells were harvested, RNA was isolated and the mRNA level of iNOS was measured by RT-PCR. β-actin was used as an internal control. The representative gels were presented (A). The values of iNOS expression were normalized to β-actin and expressed as the mean ± SD of triplicate measurements (B). iNOS activity of cell lysates was detected by the chromatometry method (C). NO concentration in cell culture supernatants was detected using NO nitrate reductase method (D). *<i>P</i><0.05, **<i>P</i><0.01.</p

Effect of TNF-α and IFN-γ on iNOS gene expression and Inhibition of intracellular Cm growth in TC-1 cell.

<p>Cultured TC-1 cells were infected with Cm at MOI 2 and treated with rmTNF-α (10 ng/ml) and/or rmIFN-γ (10 ng/ml) at 2 h postinfection. The cells were harvested at 36 h after infection and RNAs were isolated. The mRNA level of iNOS was measured by RT-PCR (A). The values of iNOS expression were normalized to β-actin. The results were presented as the mean ± SD of triplicate measurements (B). Cm was isolated from infected TC-1 cells at 36 h after infection and Cm was quantified as IFUs/ml. The percentage of inhibition was calculated as described in the Materials and Methods(C). *P<0.05, ***P<0.001.</p

Inhibition of IL-17A and IFN-γ on intracellular Cm growth in TC-1 cell.

<p>TC-1 cell monolayers were infected with Cm and treated with rmIL-17A (10 or 50 ng/ml) and/or rmIFN-γ (20 ng/ml) at 2 h post-infection. Cm inclusions in the infected TC-1 cells at 36 h after infection were stained with anti-chlamydial LPS mAb. Cm was quantified as inclusion forming units (IFUs)/ml. The representative pictures of Cm inclusions in different groups were shown (A). The percentage of inhibition was calculated as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039214#s2" target="_blank">Materials and Methods</a> (B). iNOS activity of cell lysate was detected by chromatometry method (C). The results were expressed as the mean ± SD of triplicate measurements. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001.</p

Effect of IL-17A and IFN-γ on iNOS gene expression, iNOS activity and NO production at various times following Cm infection.

<p>Cultured TC-1 cells were infected with Cm at MOI 2 and treated with rmIL-17A (10 ng/ml) and/or rmIFN-γ (20 ng/ml) at 2 h post-infection. The cells were harvested at 6, 24 and 36 h after infection and RNAs were isolated. The mRNA level of iNOS was measured by RT-PCR (A). The values of iNOS expression were normalized to β-actin. The results were presented as the mean ± SD of triplicate measurements. iNOS activity of cell lysate was detected by chromatometry method (B). NO concentration in cell culture supernatants was detected using NO nitrate reductase method (C). *P<0.05, **P<0.01, ***P<0.001.</p

Inhibition of IL-17A and IFN-γ on intracellular Cm growth in cultured ex vivo murine macrophages.

<p>Peritoneal macrophages were collected from BALB/c mice following 4% thioglycollate injection. The macrophage monolayers were then inoculated with Cm for 2 hr followed by addition of different concentrations of rmIFN-γ (10 or 200 ng/ml) and/or rmIL-17A (10 ng/ml or 100 ng/ml). Aminoguanidin (AG) was added to the wells containing IFN-γ and/or IL-17A at a final concentration of 100 µM. Chlamydial inclusions in the macrophage monolayers were stained and counted and the percentage of inhibition was calculated as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039214#s2" target="_blank">Materials and Methods</a> (A). The cells were harvested and RNA was isolated. The mRNA level of iNOS was measured by RT-PCR. The values of iNOS expression in IFN-γ and IL-17A treated cells with or without AG were normalized to β-actin (B). NO concentration in cell culture supernatant was detected using NO nitrate reductase method (C). The Cm growth (IFU) in Cm-infected macrophage in absence or presence of AG was assayed (D). The results were presented as the mean ± SD of triplicate measurements. *<i>P</i><0.05. **P<0.01, ***P<0.001.</p