RAAS in diabetic retinopathy: mechanisms and therapies.

Diabetic retinopathy (DR) is a complication of diabetes with a complex pathophysiology and multiple factors involved. Recently, it has been found that the upregulation of the renin-angiotensin-aldosterone system (RAAS) leads to overexpression of angiotensin II (Ang II), which induces oxidative stress, inflammation, and angiogenesis in the retina. Therefore, RAAS may be a promising therapeutic target in DR. Notably, RAAS inhibitors are often used in the treatment of hypertension. Still, the potential role and mechanism of DR must be further studied. In this review, we discuss and summarize the pathology and potential therapeutic goals of RAAS in DR.

RAAS in diabetic retinopathy: mechanisms and therapies

ABSTRACT Diabetic retinopathy (DR) is a complication of diabetes with a complex pathophysiology and multiple factors involved. Recently, it has been found that the upregulation of the renin-angiotensin-aldosterone system (RAAS) leads to overexpression of angiotensin II (Ang II), which induces oxidative stress, inflammation, and angiogenesis in the retina. Therefore, RAAS may be a promising therapeutic target in DR. Notably, RAAS inhibitors are often used in the treatment of hypertension. Still, the potential role and mechanism of DR must be further studied. In this review, we discuss and summarize the pathology and potential therapeutic goals of RAAS in DR.

Electrochemical enhancement of high-efficiency wet removal of mercury from flue gas.

Electrochemical wet absorption composite system has an excellent potential to remove Hg0 from flue gas. In this study, ruthenium iridium titanium platinum quaternary composite electrode is used as an anode and titanium electrode is used as the cathode, and KI/I2 absorption solution is introduced into the electrocatalysis system as an electrolyte to form KI/I2 electrochemical catalytic oxidation system. The removal rate of Hg0 in flue gas can be increased to 92.3%. The effects of electrolytic voltage, current, Pt content, I2 concentration, and the ratio of KI/I2 on the removal of Hg0 were discussed. The possible free radicals in the electrochemical cathode, anode, and solution were characterized and tested by XRD, SEM, UV-Vis (detection of H2O2, ·OH, O3), and FTIR (detection of IO3-). Combined with experimental data and theoretical derivation, the mechanism of Hg0 removal from flue gas by electrochemical catalytic oxidation alloy formation wet absorption combined process was studied. The results show that the combined process, which is a promising technology can not only improve the removal efficiency of Hg0, but also realize the resource recovery of Hg0 and I2, and provide a feasibility study for the subsequent regeneration of KI/I2 absorption solution.

Brainstem auditory evoked potential in cognitive impairment in patients with type 2 diabetes mellitus.

PURPOSE: Type 2 diabetes mellitus (T2DM) can cause mild cognitive impairment (MCI) which threatens the health of patients. So the diagnosis of MCI is particularly important. It is reported that brainstem auditory evoked potential (BAEP) is a sensitive tool to detect the brainstem function in patients with T2DM. This study aimed to investigate the relationship between BAEP and MCI in patients with T2DM. METHODS: A total of 244 T2DM patients with normal hearing, including 117 normal cognition patients and 127 MCI patients, were recruited in this cross-sectional study. Each subject underwent the BAEP examination. The diagnosis of MCI was based on the diagnostic guideline developed by the National Institute on Aging-Alzheimer's Association workgroups. The Montreal Cognitive Assessment (MoCA) was used to assess the cognitive function of the subjects. RESULTS: Compared with the normal cognition group, the patients in the MCI group had longer latencies of waves III and V and interpeak latencies (IPL) I-V in both ears (P < 0.05). The significant negative correlations were found between the latencies of waves III, V, IPL I-V, and MoCA score in both ears (P < 0.05). Logistic regression showed that the prolongations of latunits of waves III and V and IPL I-V in both ears were still associated with MCI after adjustment for mixed factors (P < 0.05). CONCLUSION: These results indicate abnormal auditory pathway in brainstem of T2DM patients with MCI. BAEP may contribute to the clinical diagnosis of MCI in patients with T2DM.

Association Between Lipid Accumulation Product and Mild Cognitive Impairment in Patients with Type 2 Diabetes.

BACKGROUND: Diabetes may increase the risk of conversion of mild cognitive impairment (MCI) to dementia. Lipid accumulation product (LAP), an index of visceral obesity, has been shown to be a powerful predictor of insulin resistance and type 2 diabetes (T2D). However, little attention has been paid to the relationship between LAP and MCI in T2D. OBJECTIVE: We aimed to investigate the association between the LAP index and MCI in patients with T2D. METHODS: In total, 220 hospitalized patients with T2D, including 113 MCI patients and 107 patients with normal cognition, were enrolled in this cross-sectional study. We collected demographic, anthropometric, and biochemical data on each subject. The LAP index was calculated according to the following formulas: [waist circumference (WC) (cm) - 65]×triglyceride (TG) (mmol/L) for males and [WC (cm) - 58] ×TG (mmol/L) for females. RESULTS: Compared with patients with normal cognition, MCI patients were older and had a higher LAP index, WC, body mass index, and glycosylated hemoglobin A1c level, as well as a lower Montreal Cognitive Assessment score and education level (p < 0.05). After adjusting for confounding factors, LAP index was associated with MCI (OR = 1.047, 95% CI = 1.031-1.063, p < 0.01). The area under the ROC curve (AUC) for the LAP index was higher than that for WC and BMI. CONCLUSION: A high LAP index is associated with an increased risk of MCI in T2D patients. The LAP index appears to be a good indicator of risk of MCI in patients with T2D.

Potential roles of Glucagon-like peptide-1 and its analogues in cognitive impairment associated with type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) is a global disease that poses a significant threat to public health. The incidence of both diabetes and dementia has increased simultaneously. Researchers have found that a large proportion of dementia patients have T2DM. In recent years, increasing evidence has demonstrated a link between cognitive decline and T2DM. Although the exact pathogenesis of cognitive impairment in T2DM is still unknown, current studies suggest that hyperglycemia, cerebrovascular disease, brain insulin resistance, and changes in γ-aminobutyric acid (GABAergic) neurons may mediate the association between T2DM and cognitive impairment. These potential mechanisms may become targets for the treatment of cognitive disorders in patients with T2DM. Glucagon-like peptide-1 (GLP-1), a widely used anti-diabetic drug, has been shown to not only effectively lower blood glucose but also improve neurological function. Previous research has confirmed that GLP-1 and its analogues are effective in the treatment of cognitive impairment in patients with T2DM. This review describes current evidence on the mechanisms underlying the association between T2DM and cognitive impairment. In particular, this review focuses on recent advances in GLP-1 and its analogues for the treatment of T2DM-related cognitive impairment.

Hypoxia-inducible factor-1α: A promising therapeutic target for vasculopathy in diabetic retinopathy.

Diabetic retinopathy (DR) is a serious condition that can cause blindness in diabetic patients. It is a neurovascular disease, but the pathogenesis leading to the onset of this disease is still not completely understood. However, hypoxia with subsequent neovascularization is a characteristic phenomenon observed with DR. Cellular response to hypoxia is mediated by the transcriptional regulator hypoxia-inducible factor (HIF). Long-term research has shown that one isotype of HIF, HIF-1α, may play a pivotal role under hypoxic conditions, and an increasing number of studies have shown that HIF-1α and its target genes contribute to retinal neovascularization. Therefore, targeting HIF-1α may lead to more effective DR treatments. This review describes the possible mechanisms of HIF-1α in neovascularization of DR. Furthermore, various inhibitors of HIF-1α that may have viable potential in the treatment of DR are also discussed.

High Serum Neuron-Specific Enolase Level Is Associated with Mild Cognitive Impairment in Patients with Diabetic Retinopathy.

PURPOSE: Diabetic retinopathy (DR) can increase the risk of mild cognitive impairment (MCI), which has been confirmed by previous researches. With the frequent occurrence of MCI in patients with DR, the early detection of MCI has become a research hot-spot. The aim of this study was to investigate the relationship between neuron-specific enolase (NSE) and MCI in patients with DR. PATIENTS AND METHODS: A total of 124 patients with DR, including 56 MCI patients and 68 normal cognition patients, were recruited in this cross-sectional study. The demographic and clinical data of patients were collected through questionnaires. Serum NSE was measured using electrochemiluminescence immunoassay. The Minimum Mental State Examination (MMSE) scale was used to evaluate the cognitive function of the participants. RESULTS: Compared with the normal cognition group, serum NSE levels and HbA1c levels in the MCI group were higher, while MMSE scores and educational level were lower (P<0.05). Serum NSE levels were significantly negatively correlated with MMSE total score, attention and calculation score, and language score (P<0.05). After adjusting for confounding factors, serum NSE still increased the MCI risk in DR patients (OR:1.606, 95CI%:1.264-2.041, P<0.001). The areas under the receiver operating characteristics (ROC) curves (AUC) of the crude model and the adjusted model were 0.75 and 0.73, respectively. CONCLUSION: A high serum NSE level is an independent risk factor for MCI in DR patients. In addition, serum NSE is expected to be a potential biomarker in DR patients with MCI.

MicroRNAs: Potential Targets in Diabetic Retinopathy.

Diabetic retinopathy (DR), a serious microvascular complication of diabetes, is a leading cause of blindness in adults. The pathogenesis of DR involves a variety of tissues and complex mechanisms, such as inflammation, oxidative stress, optic neurodegeneration, and autophagy. Nowadays, microRNAs (miRNAs), a novel group of non-coding small RNAs, have been extensively studied and recognized to play a key role in the pathogenesis of DR through aforementioned pathways. Furthermore, some miRNAs have been proposed as biomarkers that may be utilized to screen for DR. Also, miRNAs are a new therapy for DR. In this review, we summarize several miRNAs and, their roles in the pathogenesis of DR. miRNAs, as potential pharmacological targets for the diabetic retinopathy, may provide new insights for the treatment of DR.

A new research hot spot: The role of NLRP3 inflammasome activation, a key step in pyroptosis, in diabetes and diabetic complications.

Pyroptosis is a form of cell death mediated by gasdermin D (GSDMD); it is characterised by NLRP3 inflammasome activation, caspase activation, cell membrane pore formation, and the release of interleukin-1ß and interleukin-18. NLRP3 inflammasome activation plays a central role in pyroptosis. Recent research has suggested that NLRP3 inflammasome activation may be involved in the occurrence and development of diabetes mellitus and its associated complications. This finding provided the impetus for us to clarify the significance of pyroptosis in diabetes. In this review, we summarise the current understanding of the molecular mechanisms involved in pyroptosis, as well as recent advances in the role of NLRP3 inflammasome activation and pyroptosis in the development of diabetes and diabetic complications.

Evaluation of Xpert MTB/RIF for the Diagnosis of Extrapulmonary Tuberculosis in China.

We evaluate the performance of Xpert MTB/RIF for the diagnosis of extrapulmonary tuberculosis (EPTB) in China. The performance of Xpert was evaluated compared to the composite reference standard (CRS), drug susceptibility testing (DST), and imaging examination. The overall sensitivity and specificity of Xpert were 64.1% (195/304) and 100% (24/24), respectively, using CRS as the gold standard. The sensitivity was significantly higher than that of culture for pus (P<0.05). The proportion of EPTB-positive cases diagnosed by imaging was two times more than that diagnosed using Xpert; however, 6 out of 19 cases may have been overdiagnosed by imaging. Compared to phenotypic DST, the sensitivity and specificity of Xpert were 80% (12/15) and 100% (75/75), respectively. Considering its high sensitivity and specificity, Xpert MTB/RIF may be used as a rapid initial test for EPTB diagnosis, and may also support a quicker decision on the treatment regimen. The combination of imaging and Xpert testing could provide high efficiency and accurate diagnosis of suspected EPTB.

[Evaluation of the reliability of thiophen-2-carboxylic acid hydrazine result for the identification of Mycobacterium bovis].

OBJECTIVE: To evaluate the reliability of thiophen-2-carboxylic acid hydrazine (TCH) test for identification of Mycobacterium bovis (M. bovis). METHODS: A total of 4069 clinically isolated strains were identified by P-Nitrobenzoic acid medium (500 mg/L) and TCH medium (5 mg/L). Mycobacterium tuberculosis (M. tuberculosis) complex strains susceptible to 5 mg/L TCH were further tested for susceptibility to 2 mg/L TCH. Spacer oligonucleotide typing (spoligotyping) and multi-loci PCR were also performed to identify TCH susceptible strains. RESULTS: Among the 4069 isolated strains there were 3929 strains belonging to M. tuberculosis complex (MTBC) of which 245 were susceptible to 5 mg/L TCH. Of these 245 strains, 20 were also susceptible to 2 mg/L TCH, while only 1 strain was identified to be M. bovis by both spoligotyping and multi-loci PCR. CONCLUSION: TCH susceptibility test (either 5 mg/L or 2 mg/L) is not a reliable method for identification of M. bovis.

[Characteristics of soil mineralizable carbon pool in natural restoration process of Karst forest vegetation].

By the method of taking space instead of time, an incubation test was conducted to study the characteristics of soil mineralizable carbon pool during the natural restoration of Karst forest vegetation in Maolan Nature Reserve, Guizhou Province of Southwest China. It was observed that the contents of soil total organic carbon (TOC) and mineralizable carbon (MC) as well as the carbon mineralization rate decreased with increasing soil depth but increased with the process of vegetation restoration. The amount of cumulative released carbon and the carbon release rate increased with the process of restoration, but the release rate decreased with increasing incubation time. The soil MC/TOC increased with the restoration process but had less change with increasing soil depth, while the qCO2 decreased with increasing soil depth and through the process of restoration. The soil MC had a negative correlation with the existing litter amount (r = -0.796) but positive correlation with the mass loss rate of the litter decomposition (r = 0.924). Soil habitat changed from strong interference at early stages to relative stability at late stages, and soil carbon sequestration changed from small capacity and strong potential at early stages to large capacity and weak potential at late stages.

The arcuate nucleus of the C57BL/6J mouse hindbrain is a displaced part of the inferior olive.

The arcuate nucleus is a prominent cell group in the human hindbrain, characterized by its position on the pial surface of the pyramid. It is considered to be a precerebellar nucleus and has been implicated in the pathology of several disorders of respiration. An arcuate nucleus has not been convincingly demonstrated in other mammals, but we have found a similarly positioned nucleus in the C57BL/6J mouse. The mouse arcuate nucleus consists of a variable group of neurons lying on the pial surface of the pyramid. The nucleus is continuous with the ventrolateral part of the principal nucleus of the inferior olive and both groups are calbindin positive. At first we thought that this mouse nucleus was homologous with the human arcuate nucleus, but we have discovered that the neurons of the human nucleus are calbindin negative, and are therefore not olivary in nature. We have compared the mouse arcuate neurons with those of the inferior olive in terms of molecular markers and cerebellar projection. The neurons of the arcuate nucleus and of the inferior olive share three major characteristics: they both contain neurons utilizing glutamate, serotonin or acetylcholine as neurotransmitters; they both project to the contralateral cerebellum, and they both express a number of genes not present in the major mossy fiber issuing precerebellar nuclei. Most importantly, both cell groups express calbindin in an area of the ventral hindbrain almost completely devoid of calbindin-positive cells. We conclude that the neurons of the hindbrain mouse arcuate nucleus are a displaced part of the inferior olive, possibly separated by the caudal growth of the pyramidal tract during development. The arcuate nucleus reported in the C57BL/6J mouse can therefore be regarded as a subgroup of the rostral inferior olive, closely allied with the ventral tier of the principal nucleus.

[Changes of foliar delta13C value of Quercus fabric in different root underground habitat types in Karst area].

Selecting the dominant tree species Quercus fabric in three root underground habitat types (the dolomites of low oblique occurrence with multilayer space, middle oblique occurrence with multilayer space, and high oblique occurrence with multilayer space) in Karst area as test object, this paper studied the foliar delta13C value and its correlations with habitat soil conditions, and the plant water use efficiency. There existed remarkable differences in the foliar delta13C value of Q. fabric among the three habitat types, being decreased in the order of low oblique occurrence with multilayer space type (-26.35 per thousand) > high oblique occurrence with multilayer space type (-26.66 per thousand) > middle oblique occurrence with multilayer space type (-27.07 per thousand). Accordingly, the plant water use efficiency decreased in the same order. The foliar delta13C value had significant correlation with habitat soil moisture content, but less correlation with habitat soil elements contents. The delta13C value increased with the decrease of soil moisture content and soil fertility.

[Characteristics of soil microbial biomass carbon and soil water soluble organic carbon in the process of natural restoration of Karst forest].

By the method of taking space instead of time, an incubation test was conducted to study the characteristics of soil microbial biomass carbon and water soluble organic carbon in the process of natural restoration of Karst forest in Maolan Nature Reserve, Guizhou Province of Southwest China. The soil microbial biomass carbon content and soil basal respiration decreased with increasing soil depth but increased with the process of the natural restoration, soil microbial quotient increased with increasing soil depth and with the process of restoration, and soil water soluble organic carbon content decreased with increasing soil depth. In the process of the natural restoration, surface soil water soluble organic carbon content increased, while sublayer soil water soluble organic carbon content decreased after an initial increase. The ratio of soil water soluble organic carbon to total soil organic carbon increased with increasing soil depth but decreased with the process of restoration. Soil quality increased with the process of restoration. Also, the quality and quantity of soil organic carbon increased with the process of restoration, in which, soil microbial biomass carbon content had the greatest change, while soil water soluble organic carbon content had less change.

[Relationship between Beijing genotypes of Mycobacterium tuberculosis and drug-resistant phenotypes].

OBJECTIVE: To investigate the distribution of the Beijing genotypes of Mycobacterium tuberculosis (M. tuberculosis) and the relationships between Beijing genotype strains and drug-resistant phenotypes in China. METHODS: Clinical isolates were collected during a 9-month research period from April to December in 2008 in six geographic regions of China. One isolate that had been biochemically confirmed to be a member of the M. tuberculosis complex was collected from each patient. The demographic data of the patients (eg. sex, age, and history of tuberculosis) as well as the drug resistance patterns and sources of the clinical isolates were collected. Drug susceptibility testing was performed using proportion method. Beijing genotypes of M. tuberculosis were identified by spacer oligonucleotide typing or insertion of IS6110 in the genomic dnaA-dnaN locus. RESULTS: Among the 410 M. tuberculosis clinical isolates, 67.1% (275/410) isolates were Beijing genotypes of M. tuberculosis. Significantly larger proportions of tuberculosis patients were infected with Beijing genotypes in the northeastern regions of China than that of in the central-western regions (chi2 = 20.50, P = 0.000). No significant associations were found either between Beijing genotype strains and patients' age, sex, or treatment history. Multidrug-resistant isolates and rifampin-resistant isolates were more common among Beijing genotype strains than among non-Beijing strains (P = 0.002, P = 0.005). CONCLUSIONS: About two third of the clinical isolates of M. tuberculosis in China are Beijing genotypes. Beijing genotype strains are not correlated with patients' age, sex, treatment history. People living in the northeastern regions of China are more susceptible to Beijing genotypes than those living in the central-western of China. Beijing genotype strains tend to be rifampin-resistant or multidrug-resistant.

[1, 25-dihydroxyvitamin D3 promotes the apoptosis of inflammatory cells in acute experimental autoimmune encephalomyelitis: experiment with rats].

OBJECTIVE: To investigate the mechanism of rapid amelioration of the pathological changes in experimental allergic encephalomyelitis (EAE) by 1, 25-dihydroxyvitamin D(3) [1, 25-(OH)(2)D(3)]. METHODS: Forty Lewis rats were immunized with myelin basic protein in complete Freud's adjuvant so as to establish ESE animal models and then randomly divided into 4 equal groups: prevention group, fed with 1, 25-(OH)(2)D(3) since day 0 for 10 days, prevention-control group fed with peanut oil for 10 days, treatment group fed with 1, 25-(OH)(2)D(3) since the appearance of EAE symptoms (generally since day 10 or 11), and treatment-control group fed with peanut oil since the appearance of EAE symptoms. The clinical symptoms were scored since immunization till day 12 when the clinical symptoms reached the maximum level. The rats were sacrificed 13 days after sensitization with their brains and spinal cords taken out to undergo pathological examination, in situ TUNEL staining for detecting apoptotic cells, and semiquantitative immunohistochemical analysis to detect the inducible NO synthase (iNOS), FasL, and tumor growth factor (TGF)-beta 1, that might involve in the signal pathway of apoptosis. Peripheral blood samples were collected to isolate mononuclear cells (MNCs). The content of nitrite in the supernatant of MNC culture was evaluated. RESULTS: The scores of clinical symptoms and the pathological changes of both the prevention and treatment groups decreased conspicuously and were significantly lower than their respective control groups (both P < 0.01). In contrast, the apoptosis indexes of the 2 1, 25-(OH)(2)D(3) administration groups were significantly higher than those of the control groups (all P < 0.01). The TUNEL positive cell rates in the brain and spinal cord of the treatment and prevention groups were all significantly higher than those of their corresponding control groups (P < 0.05, P < 0.01). The numbers of iNOS positive cells in the treatment and prevention groups were both lower than those of their corresponding control groups, which was in accord with the improvement of clinical signs and tissue lesions. The levels of nitrite in the supernatant of MNC culture of the treatment and prevention groups were higher than those of their corresponding control groups, but not significantly. CONCLUSION: Administration of 1, 25-(OH)(2)D(3) rapidly ameliorates EAE symptoms by promoting the apoptosis of inflammatory cells. The elimination of infiltrating immune cells which reverses the pathological changes in central nervous system is associated with a favorable microenvironment provided by 1, 25-(OH)(2)D(3), such as decreasing of iNOS.

[In vitro study on cross resistance of rifampin and rifapentine for Mycobacterium tuberculosis].

OBJECTIVE: To study the bactericidal effect of rifapentine and its cross-resistance with rifampin for Mycobacterium tuberculosis and to determine the critical concentration of rifapentine for laboratory drug susceptibility test and therefore to provide the laboratory data for using rifapentine in the treatment of tuberculosis, particularly rifampin resistant tuberculosis. METHODS: We detected the minimal inhibitory concentrations (MICs) of rifampin and rifapentine to H(37) Rv and isolated strains of rifampin susceptible and resistant Mycobacterium tuberculosis by using Middlebrook 7H9, Sauton and Lowenstein-Jensen media. RESULTS: The MICs of rifampin were > or = 0.32 microg/ml for 80% of the 19 rifampin susceptible strains on Middlebrook 7H9 and the MICs of rifapentine ranged from 0.02 microg/ml to 0.32 microg/ml for most of the strains (84%). The MICs of rifapentine were 2 - 4 times lower than those of rifampin to H(37) Rv and most clinical isolates. The rifapentine susceptible isolates were mostly separated from resistant strains at MICs 5-10 microg/ml. CONCLUSIONS: Our results demonstrate the cross resistance of rifampin and rifapentine and the stronger bactericidal potency of rifapentine than rifampin. Some rifampin resistant strains still show susceptibility to rifapentine, which suggests rifapentine may be effective in the treatment of rifampin resistant tuberculosis. Our results also determined a critical resistant concentration of rifapentine for routine drug susceptibility test.

[Mechanism of cellular immunity accommodation in prophylactic effects of nasal tolerance with dual analogue on experimental autoimmune myasthenia gravis in Lewis rats].

OBJECTIVE: To study the mechanism of prophylactic effects of nasal tolerance with a dual analogue (Lys262-Ala207) on experimental autoimmune myasthenia gravis (EAMG). METHODS: Clinical and immunological changes were observed in Lewis rats administered with dual analogue Lys262-Ala207 nasally, to compare the effects between the rats with predetermined dosage of Lys262-Ala207 and control peptides at two different time points, before the day (Group A or C) or on the day (Group B or D) of immunization with acetylcholine receptor (AChR) in complete Freud's adjuvant for 10 consecutive days. The clinical scores was evaluated for 50 days post immunization. Numbers of MNC expressing IFN-gamma, IL-4 or IL-10 and CD4+ and/or CD25+ from lymph nodes were enumerated by flow cytometry. Proliferative response, expressed as stimulation index (SI), was suppressed in response to antigen-specific stimulation in the rats receiving dual analogue, as compared with the rats receiving saline buffer only. RESULTS: Group A and group B of Lewis rats developed EAMG with reduced severity, as compared to the control groups. Number of cells synthesizing IFN-gamma, IL-4 or IL-10 decreased, whereas numbers of CD4+CD25+ cells increased in group A and B than those in the control groups. Proliferative response was suppressed in response to antigen-specific stimulations in the rats receiving dual analogue Lys262-Ala207. CONCLUSIONS: Nasal administration with a dual analogue Lys262-Ala207 at two different time points, before the day and on the day of immunization, could delay symptoms of muscular weakness in EAMG rats, which was associated with suppression of immune function in AChR antigen-specific T cells and lay a scientific foundation for treatment of human MG with nasal dual analogue.