Negative ANA-IIF in SLE patients: what is beyond?

The antinuclear antibody (ANA) test has high sensitivity in diagnosing and classifying systemic lupus erythematosus (SLE). OBJECTIVES: To describe the immunological pattern of SLE patients through investigating specific antinuclear autoantibodies by enzyme dot immunoassay and studying their frequency in both positive and negative ANA indirect immunofluorescence assay (IIF) cases. METHODS: In a cross-sectional study, blood samples from 393 newly diagnosed SLE patients were analyzed using (IIF) on HEp-2 cells and ANA dot immunoassay by automated enzyme immunoassay (EIA) to detect 19 antibodies. RESULTS: Ninety-one percent of the patients are females; their mean age was 37 ± 12.28. Antinuclear antibody (ANA) was detected by IIF in 82.4% of cases, with 181 (46.1%) speckled and 167 (42.4%) homogeneous ANA patterns. The majority of patients (96%) demonstrated autoantibodies via EIA. Among the ANA-IIF-negative patients, 97.2% demonstrated autoantibodies. There was a significant difference in the frequency of certain autoantibodies between SLE patients with negative and positive ANA-IIF (1.44 0.73, 3.12 2.09, p = 0.00) respectively. CONCLUSION: The results of analyzing 19 autoantibodies with the ANA staining pattern increased the significance of analyzing the immune profile even if IIF is negative when clinical symptoms strongly suggest SLE diagnosis. Certain autoantibodies may evade staining by the IFA approach while they are present in the patient's serum, and they may not be detected by the ANA EIA profile if it does not contain that antigenic substrate. Key Points • Indirect immunofluorescence on Hep-2 is the conventional method for ANA detection and is regarded as the "gold standard" for testing in clinical practice for SLE. • In our study, ANA profile dot enzyme immunoassay (EIA)-based test was performed to evaluate 19 autoantibodies in SLE patients either positive or negative for ANA-IIF. • The presence of anti-dsDNA with ANA-IIF-negative serum in 32.4% of SLE patients provides evidence that not all anti-dsDNA antibodies are identified on standard HEp-2 substrates. • certain autoantibodies can evade staining by the ANA-IIF method despite being present in the SLE patient's blood; this supports the ANA profile enzyme dot immunoassay as a more sensitive test.

Immunoglobulin G antibody immune response profile following infection with SARS-CoV-2 in COVID-19 Egyptian patients.

This study aimed to report the dynamic profile of IgG-specific antibodies to SARS-CoV-2 infection for 6 months after infection. We conducted a prospective study, recruited 33 recently confirmed covid -19 patients and collected 6 samples from each patient. The first samples were collected one month from the start of symptoms and subsequent samples collected at 30 days interval. We measured the IgG by chemiluminescent immunoassay (CLIA). According to the disease severity, patients were categorized as asymptomatic 4 (12.1%), mild 14 (42,4%), moderate 9 (27.3%), and severe 6 (18.2%). Patients were 12 (35.3%) females and 21 (64.7%) males. The mean IgG levels maintained a high level till the second month (92.81 ± 110.15 AU/ml) from the onset of symptoms followed by a gradual decrease till the sixth month after infection (17.42 ± 22.61 AU/ml). The patients with severe symptoms significantly exhibited the highest IgG levels, reached the highest level (mean=237.44 ± 164.13 AU/ml) at the second month. While the lowest levels were detected among the asymptomatic patients (mean= 3.04 ± 2.94 AU/ml) at the second month. Older age correlated with higher IgG antibody level (r= 0.350 p=0.046); however, sex was not related to IgG level. In conclusion, Symptomatic COVID-19 disease is followed by protective immunity for more than 6 months. Immunity in asymptomatic patients is low and fades rapidly than symptomatic cases. Patients with severe disease had significantly higher IgG levels compared to mild, moderate, or asymptomatic patients.

Significant Interrelations among Serum Annexin A1, Soluble Receptor for Advanced Glycation End Products (sRAGE) and rs2070600 in Chronic Obstructive Pulmonary Disease.

Chronic obstructive pulmonary disease (COPD) is a major cause of death and morbidity; it may be accompanied by oxidative stress and inflammation with or without underlying genetic etiology. Finding circulating biomarkers for COPD that can help early diagnosis and predict exacerbation and association with respiratory functions has been challenging. There were 40 healthy participants and 60 COPD patients in this research. The rs2070600 gene variant was examined by PCR-RFLP. Circulating sRAGE and annexin A1 levels were determined by ELISA. GSH and MDA were determined by spectrophotometry. In COPD patients, sRAGE serum levels were substantially lower, but conversely, annexin A1 levels were much greater than in controls. The rs2070600 gene polymorphism's strong association with COPD was demonstrated by genotyping and allelic frequency distribution. The GA genotype was most distributed in COPD, and it was strongly linked to lower serum sRAGE levels. The interrelation between annexin A1, sRAGE, and COPD could be explained through effects on inflammatory mediators' pathways. The rs2070600 gene polymorphism was found to significantly enhance the risk of COPD. Serum sRAGE and annexin A1 may be considered potential diagnostic tools for COPD. Through impacts on GSH and MDA levels that alter the release of inflammatory factors and, therefore, lung damage, it is possible to explain the relationship between annexin A1, sRAGE, and COPD.

Assessment of Ferula hermonis Boiss fertility effects in immature female rats supported by quantification of ferutinin via HPLC and molecular docking.

ETHNOPHARMACOLOGICAL RELEVANCE: Ferula hermonis is a small shrub renowned for its aphrodisiac abilities. Middle East herbalists have utilized Ferula hermonis seed and root as an aphrodisiac folk medicine to treat women's frigidity and male erectile and sexual dysfunction. AIM OF THE STUDY: Assessment of follicle-stimulating hormone-like (FSH), luteinizing hormone-like (LH), and estrogenic activities of the methanolic extract (ME) of the roots of Ferula hermonis on female reproductive function. MATERIALS AND METHODS: The methanolic extract was prepared from the root of F. hermonis and studied at dose level 6 mg/kg in immature female rats for FSH-like, LH-like, and estrogenic activities. These activities were determined by analyzing gross anatomical features, relative organ weight, and serum level of FSH, LH, progesterone and estrogen hormones, and histopathological characteristics. Quantification of the main phytoestrogenic component ferutinin carried out by HPLC. In addition, molecular docking for the binding affinity of ferutinin inside active sites of both estrogen receptor alpha (ERα) and FSH receptor (FSHR) was performed to predict the potential role of ferutinin in regulating the female reproductive process. RESULTS: Ferula hermonis (ME) showed potent FSH-like, LH-like activities and moderate estrogenic effect at the dose of 6 mg/kg. The content of ferutinin in F. hermonis was estimated to be 92 ± 1.33 mg/g of the methanolic extract. Molecular docking of ferutinin with ERα and FSHR displayed strong interaction with target proteins. CONCLUSIONS: Based on results, it can be concluded that Ferula hermonis can be considered as a suitable female fertility improving agent.

Association of anti-neutrophil cytoplasmic antibody in ischemic stroke Egyptian patients with hepatitis C virus.

Some studies reported a high prevalence of ischemic stroke in hepatitis C virus patients, other several studies have suggested that hepatitis C virus (HCV) may act as a trigger for autoimmune diseases and autoantibodies including Anti-Neutrophil Cytoplasmic Antibody (ANCA) which predispose to vasculitis. Because vasculitis is a risk factor for ischemic stroke, we investigated the association of the hepatitis C virus with ANCA in first-ever ischemic stroke patients. This study included 67 Egyptian patients with first-ever ischemic stroke. These patients were clinically examined and investigated for HCV infection by chemiluminescence & Real Time-PCR, and ANCA antibodies by ELISA. Forty-two patients (62.7%) had HCV infection. Twenty-nine (43.2%) of them were cytoplasmic- Antineutrophil Cytoplasmic Antibodies (c-ANCA) positive, while none was perinuclear- Antineutrophil Cytoplasmic Antibodies (p-ANCA) positive. Comparison between c-ANCA positive and ANCA negative patients showed that 82.8% and 47.4% had anti-HCV antibody, respectively, with P-value 0.003. The c-ANCA level correlated significantly with age, and HCV antibody level. No statistically significant difference was found in both the consciousness and stroke severity between the negative and positive c- ANCA patients. However, patients with positive c-ANCA had smaller and multiple cerebral infarctions with P-value 0.002 and 0.01 respectively. Multiple regression analysis showed that the number and size of cerebral infarctions were independent predictors of c-ANCA positivity with P value 0.02, and 0.03 respectively. In conclusion, c-ANCA level correlates with HCV antibody and may predispose to ischemic stroke by a possible ANCA associated vasculitis.

Impact of travel by walk and road on testicular hormones, oxidants, traces minerals, and acute phase response biomarkers of dromedary camels.

This study aimed to compare the effect of truck transport and walk travel on testicular hormones, oxidants, antioxidants and acute-phase responses of camels' walked from Sudan to the Egyptian quarantine and were transported from the quarantine to the slaughterhouses by trucks. Blood samples were collected from walked camels (N ≤ 30) just arrived at the quarantine (Walk), unloaded (N ≤ 12) from the truck (Truck), and control camels (N ≤ 20). Animals were statistically categorized into Walk travel, Truck transport, and Control, then Total travel (Walk + truck transport) was compared to control. Haptoglobin, fibrinogen, superoxide dismutase (SOD), glutathione peroxidase (GPx), nitric oxide (NO), ascorbic acid, glucose, cholesterol, testosterone, estradiol, iron, copper, ALT, AST, alkaline phosphatase (ALP), total proteins, albumin, and creatinine were measured. Results showed that the travel by walk and truck increased haptoglobin (P ≤ 0.0001), fibrinogen (P < 0.05), ALT (P < 0.05), and creatinine (P ≤ 0.0001) but decreased NO (P ≤ 0.0001), albumin (P < 0.05), Ascorbic acid (P < 0.05), testosterone (P ≤ 0.0001), ALP (P < 0.0001), and glucose (P ≤ 0.0001). The declined NO (P ≤ 0.0001), Ascorbic acid (P ≤ 0.0001), iron (P ≤ 0.005), copper (P ≤ 0.023), cholesterol (P > 0.05), total proteins (P ≤ 0.0001), albumin (P ≤ 0.018), globulins (P ≤ 0.001), with increased haptoglobin (P ≤ 0.0001), AST (P ≤ 0.0001), ALP (P ≤ 0.0001), and testosterone (P ≤ 0.0001) was evident in camels transported by truck compared to walk transport. In conclusion, transport enhanced the acute phase proteins, retarded kidney function, antioxidant status, and energy but truck produced a significant acute-phase response and adversely affected the oxidant-antioxidant balance, destructed proteins kidney, and liver functions than the long travel by walk.

Assessment of Coronavirus in the Conjunctival Tears and Secretions in Patients with SARS-CoV-2 Infection in Sohag Province, Egypt.

PURPOSE: To assess SARS-CoV-2 virus in conjunctival tears and secretions of positive confirmed COVID-19 patients. METHODS: A case series study that included 28 positive COVID-19 patients confirmed with nasopharyngeal swab in the period 18-28 May 2020 at Sohag Tropical Medicine Hospital. Tears and conjunctival secretions of these confirmed positive cases were collected with disposable sampling swabs at interval of 3 days after admission due to respiratory symptoms. They were examined for the presence of SARS-CoV-2 by reverse transcription-polymerase chain reaction (RT-PCR) assay. RESULTS: Thirteen (46.43%) patients were stable, 4 (14.28%) patients suffered from dyspnea, 3 (10.72%) patients suffered from high fever, 5 (17.85%) patients suffered from cough, and 3 (10.72%) patients were on mechanical ventilation. Ten (35.71%) patients suffered from conjunctivitis. Tear and conjunctival swabs were positive in 8 (28.57%) patients, while other patients' swabs were negative (71.43%). Out of 10 patients with conjunctival manifestations, 3 patients had SARS-CoV-2 in their conjunctiva using (RT-PCR) test. Out of the 18 patients with no conjunctival manifestations, 5 patients had positive SARS-CoV-2 in their conjunctiva using (RT-PCR) test. CONCLUSION: The SARS-CoV-2 virus could be found in tears and conjunctival secretions in SARS-CoV-2 patients with or without conjunctivitis.

Effect of exogenous progesterone treatment on ovarian steroid hormones and oxidant and antioxidant biomarkers during peak and low breeding seasons in dromedary she-camel.

BACKGROUND: Research about the effects of progesterone (P4) and the relationship of P4 to oxidative stress has been achieved in ruminants but not enough in camels. AIM: This study evaluated the effect of exogenous P4 hormone using CIDR for 7 days on blood concentrations of steroid hormones and oxidative status of dromedary she-camels during peak and low breeding seasons. MATERIALS AND METHODS: The present work was conducted on ten dark dromedary she-camels which were synchronized using a controlled internal drug release (CIDR) for 7 days as a reproductive management tool during peak breeding (November-April) and low breeding season (May-October). The blood samples were collected each other day from CIDR insertion until the end of experiment 5 days after the removal of CIDR. Camels were examined for P4, estradiol (E2), and testosterone (T) as well as malondialdehyde (MDA) as indicator of lipid peroxidation and nitric oxide, superoxide dismutase (SOD), and glutathione-S-transferase as antioxidant markers. RESULTS: Results revealed that P4 was higher during peak breeding season than low breeding season. While the levels of P4 increased during CIDR insertion and declined at CIDR removal and thereafter during breeding season, its concentrations declined after CIDR application during the non-breeding season. On the other hand, blood E2 and testosterone levels decreased after CIDR insertion in both high and low breeding seasons with higher serum E2 concentrations during the peak than the low breeding season. MDA concentrations and SOD activities were significantly (p<0.05) high on day 3 after CIDR insertion during the breeding and non-breeding seasons. During both the seasons, GSH levels decreased after CIDR removal in camels. However, MDA was lower during non-breeding season than high breeding season with no seasonal effect on SOD activity. CONCLUSION: Exogenous P4 treatment through CIDR in dromedary camels could be more efficient during breeding season than non-breeding season, and effects on circulating oxidant/antioxidant biomarkers and their return to normal levels might refer to the adaptation of camels to CIDR by modulating their oxidant and antioxidant levels.

Ovarian hormones and antioxidant biomarkers in dromedary camels synchronized with new and re-used controlled intravaginal drug release (CIDR)/GPG (Ovsynch) program during breeding season.

This study aimed to investigate the effect of CIDR, re-used wCIDR, and Ovsynch protocols for the synchronization of follicular waves on ovarian hormones, oxidative stress, and antioxidant biomarkers during the breeding season. Dromedary camels (N = 18) were divided into three equal groups. The first group received CIDR. The second group received previously used wCIDR after thorough cleaning and disinfection. The third group was subjected to GPG protocol. Progesterone (P4), estradiol (E2) l, total antioxidant capacity (TAC), catalase (CAT), lipid peroxide product (MDA), nitric oxide (NO), and glutathione reduced (GSH) were measured. Days during CIDR affected P4 (P = 0.0001), E2 (P = 0.047), TAC (P = 0.01), NO (P = 0.028), and GSH (P = 0.005). Days during re-used wCIDR effected P4, TAC, CAT, NO, GSH, and MDA (P ≤ 0.001). Days during GPG effected P4, E2, TAC, GSH (P = 0.0001), MDA (P = 0.036), and NO (P = 0.02). CIDR-treated camels had high P4 (P = 0.0001), E2 (P = 0.0001), TAC (P = 0.012), and NO (P = 0.0001), with low GSH (P = 0.001), and MDA (P = 0.003). Exogenous progesterone improved ovarian hormones and the antioxidant capacity and minimized the oxidative stress than the GPG treatment and is recommended for future reproductive management of camels.

The Role of Bcl-2 and Bax as Markers of Disease Progression in Hepatitis C Virus Infected Patients.

Many apoptotic markers have been linked to hepatic cell injury in HCV-related liver diseases, and hence could be used as potential markers for early detection of the disease. The present study aimed to assess the role of apoptotic markers Bcl-2 and Bax in the pathogenesis of chronic HCV-related liver diseases. A total of 85 participants were enrolled into the study; 70 chronic HCV patients (35 non-cirrhotic and 35 cirrhotic), and 15 healthy controls. The serum levels of Bcl-2 and Bax were assayed in all participants by ELISA. Bcl-2 and Bcl-2/Bax ratio were significantly higher in non-cirrhotic patients than the cirrhotic and controls (P < 0.001). Bax was significantly higher in cirrhotic patients compared to the other groups (P < 0.001). Positive and negative correlations were found between serum Bcl-2, Bax, Bcl-2/Bax ratio and HCV viral load in non-cirrhotic and cirrhotic patients respectively. These findings provide an evidence that apoptosis is dysregulated in patients with chronic HCV.

Exploring the PDE5 H-pocket by ensemble docking and structure-based design and synthesis of novel ß-carboline derivatives.

By studying the co-crystal information of interactions between PDE5 and its inhibitors, forty new tetrahydro-ß-carbolines based-analogues were synthesized, and tested for their PDE5 inhibition. Some compounds were as active as tadalafil in inhibiting PDE5 and of better selectivity profile particularly versus PDE11A, the nature of the terminal ring and its nitrogen substituent are the main determinants of selectivity. Ensemble docking confirmed the role of H-loop closed conformer in activity versus its occluded and open forms. Conformational studies showed the effect of bulkiness of the terminal ring N-alkyl substituent on the formation of stable enzyme ligands conformers. The difference in potencies of hydantoin and piperazinedione analogues, together with the necessity of C-5/C-6 R-absolute configuration has been revealed through molecular docking.