Coinfection and cross-reaction of dengue and COVID-19: a case series analysis.

BACKGROUND: The risk of possible cross-reactions between serological tests, together with the clinical similarities between dengue fever and COVID-19, can delay diagnosis and increase the risk of both COVID-19 transmission and worsening. The present study aimed to determine the possibility of cross-reactions among rapid serological tests based on clinical symptoms. METHODS: Patients with COVID-19, confirmed by RT-PCR and clinical criteria for diagnosing dengue, were recruited consecutively between September 2020 and August 2021 and underwent rapid immunochromatographic diagnostic (RID) tests for AgNS1, IgM, and IgG. Patients who tested positive for acute-phase dengue IgM and AgNS1 underwent a follow-up test after 12-30 days for diagnostic confirmation. RESULTS: A total of 43 patients were included, 38 of whom required hospital admission, and 8 received intensive care. Seven patients tested positive on the RID tests, comprising 2 NS1 positive (coinfection), one reactive for IgM and IgG (coinfection), three reactive for IgM not confirmed (false-positive), and one reactive for IgG due to previous infection. Two of the 3 patients with coinfection died. Fever, myalgia, headache, and cough were the most common clinical symptoms, while lymphopenia was the most prevalent laboratory finding. CONCLUSIONS: Cross-reactivity was found in only three patients and coinfection in another three patients, two of whom died of severe COVID-19 manifestations.

Coinfection and cross-reaction of dengue and COVID-19: a case series analysis

ABSTRACT Background: The risk of possible cross-reactions between serological tests, together with the clinical similarities between dengue fever and COVID-19, can delay diagnosis and increase the risk of both COVID-19 transmission and worsening. The present study aimed to determine the possibility of cross-reactions among rapid serological tests based on clinical symptoms. Methods: Patients with COVID-19, confirmed by RT-PCR and clinical criteria for diagnosing dengue, were recruited consecutively between September 2020 and August 2021 and underwent rapid immunochromatographic diagnostic (RID) tests for AgNS1, IgM, and IgG. Patients who tested positive for acute-phase dengue IgM and AgNS1 underwent a follow-up test after 12-30 days for diagnostic confirmation. Results: A total of 43 patients were included, 38 of whom required hospital admission, and 8 received intensive care. Seven patients tested positive on the RID tests, comprising 2 NS1 positive (coinfection), one reactive for IgM and IgG (coinfection), three reactive for IgM not confirmed (false-positive), and one reactive for IgG due to previous infection. Two of the 3 patients with coinfection died. Fever, myalgia, headache, and cough were the most common clinical symptoms, while lymphopenia was the most prevalent laboratory finding. Conclusions: Cross-reactivity was found in only three patients and coinfection in another three patients, two of whom died of severe COVID-19 manifestations.

Thermoresponsive Hydrogel-Loading Aluminum Chloride Phthalocyanine as a Drug Release Platform for Topical Administration in Photodynamic Therapy.

Phthalocyanine aluminum chloride (Pc) is a clinically viable photosensitizer (PS) to treat skin lesions worsened by microbial infections. However, this molecule presents a high self-aggregation tendency in the biological fluid, which is an in vivo direct administration obstacle. This study proposed the use of bioadhesive and thermoresponsive hydrogels comprising triblock-type Pluronic F127 and Carbopol 934P (FCarb) as drug delivery platforms of Pc (FCarbPc)-targeting topical administration. Carbopol 934P was used to increase the F127 hydrogel adhesion on the skin. Rheological analyses showed that the Pc presented a low effect on the hydrogel matrix, changing the gelation temperature from 27.2 ± 0.1 to 28.5 ± 0.9 °C once the Pc concentration increases from zero to 1 mmol L-1. The dermatological platform showed matrix erosion effects with the release of loaded Pc micelles. The permeation studies showed the excellent potential of the FCarb platform, which allowed the partition of the PS into deeper layers of the skin. The applicability of this dermatological platform in photodynamic therapy was evaluated by the generation of reactive species which was demonstrated by chemical photodynamic efficiency assays. The low effect on cell viability and proliferation in the dark was demonstrated by in vitro assays using L929 fibroblasts. The FCarbPc fostered the inhibition of Staphylococcus aureus strain, therefore demonstrating the platform's potential in the treatment of dermatological infections of microbial nature.

Pharmacokinetics of amoxicillin in obese and nonobese subjects.

AIMS: To compare the pharmacokinetics of amoxicillin (AMX) in obese and nonobese subjects, given as single dose 875-mg tablets. METHODS: A prospective, single-centre, open-label, clinical study was carried out involving 10 nonobese and 20 obese subjects given a dose of an AMX 875-mg tablet. Serial blood samples were collected between 0 and 8 hours after administration of AMX and plasma levels were quantified by liquid chromatography-tandem mass spectrometry. The pharmacokinetic parameters (PK) were calculated by noncompartmental analysis and means of the 2 groups were compared using Student t-test. Analysis of correlation between covariates and PK was performed using Pearson's correlation coefficient. RESULTS: Ten nonobese subjects (mean age 30.6 ± 7.12 y; body mass index 21.56 ± 1.95 kg/m2 ) and 20 obese subjects (mean age 34.47 ± 7.03 y; body mass index 33.17 ± 2.38 kg/m2 ) participated in the study. Both maximum concentration (Cmax ; 12.12 ± 4.06 vs. 9.66 ± 2.93 mg/L) and area under the curve (AUC)0-inf (34.18 ± 12.94 mg.h/L vs. 26.88 ± 9.24 mg.h/L) were slightly higher in nonobese than in obese subjects, respectively, but differences were not significant. The volume of distribution (V/F) parameter was statistically significantly higher in obese compared to nonobese patients (44.20 ± 17.85 L vs. 27.57 ± 12.96 L). Statistically significant correlations were observed for several weight metrics vs. AUC, Cmax , V/F and clearance, and for creatinine clearance vs. AUC, Cmax and clearance. CONCLUSION: In obese subjects, the main altered PK was V/F as a consequence of greater body weight. This may result in antibiotic treatment failure if standard therapeutic regimens are administered.

Curcumin and silver nanoparticles carried out from polysaccharide-based hydrogels improved the photodynamic properties of curcumin through metal-enhanced singlet oxygen effect.

Colorectal cancer (CRC) has a high incidence and resistance to conventional treatments. Curcumin (CUR) is a promising natural product in the treatment of CRC with excellent in vitro results. However, its low bioavailability is a limiting factor in clinical applications. To overcome, CUR was incorporated into hydrogels constituted by chitosan (CHT) and chondroitin sulfate (CS), natural biopolymers, capable of controlled release. Hydrogels were synthesized in ionic liquids (ILs, [Hmim][HSO4]) improving the solubility of CHT and the hydrogel properties. Furthermore, CUR was combined with silver nanoparticles (AgNPs) and visible light by Photodynamic Therapy (PDT), which, through the MEO effect (Metal-Enhanced Singlet Oxygen), leads to cell death. It is highlighted the green synthesis of AgNPs using an ultrasound bath. The CHT/CS hydrogels loaded with CUR/AgNPs were properly characterized. Cellular assays showed that the hydrogels (CHT/CS) were not cytotoxic to healthy tissues. However, PDT selective illumination led to inhibition of Caco-2 human colon cancer cells by the CHT/CS/CUR-AgNPs (CC50 = 91.5 µg mL-1 of hydrogel). The cellular uptake assays showed, in addition to the therapeutic action, that the CUR can works as a diagnostic fluorescence probe (theranostic system). Finally, we highlight our commitment to work with reagents, solvents, and methodologies aiming at the principles of green chemistry.

Population pharmacokinetics of orally administrated bromopride: Focus on the absorption process.

Bromopride is a prokinetic and antiemetic drug used to treat nausea and vomiting. Although its prescription is common in Brazil, there is a lack of studies about bromopride pharmacokinetics. Therefore, the aims of this study were to investigate the population pharmacokinetics of bromopride and to evaluate the influence of covariates on its absorption. This study is a retrospective analysis of data collected from bioequivalence studies. The data was modeled using MONOLIX 2018R2. Assuming one-compartment and linear elimination, the absorption phase was evaluated with different structural models. The model of sequential first- and zero-order with combined error and exponential inter-individual variability in all parameters best described the atypical absorption profile of bromopride. Population estimates were first-order absorption rate (ka) of 0.08 h - 1, fraction of dose absorbed by first-order (Fr) of 32.60%, duration of the zero-order absorption (Tk0) of 0.88 h with latency time (Tlag) of 0.47 h, volume of distribution of 230 l and clearance of 46.80 l h - 1. Bodyweight affects Tk0, dosage form was found to correlate with Tk0 and Tlag, while gender affects Tlag. However, simulations evaluating the clinical importance of these covariates on steady-state indicated minimal changes on bromopride exposure. The mixed absorption model was reasonable to describe the absorption process of bromopride because it had the flexibility to fit multiple-peaks profile and shows good agreement with physicochemical properties of drug.

Selective Photodynamic Effects on Breast Cancer Cells Provided by p123 Pluronic®- Based Nanoparticles Modulating Hypericin Delivery.

BACKGROUND: Breast cancer is the most relevant type of cancer and the second cause of cancer- related deaths among women in general. Currently, there is no effective treatment for breast cancer although advances in its initial diagnosis and treatment are available. Therefore, the value of novel anti-tumor therapeutic modalities remains an immediate unmet need in clinical practice. Following our previous work regarding the properties of the Pluronics with different photosensitizers (PS) for photodynamic therapy (PDT), in this study we aimed to evaluate the efficacy of supersaturated hypericin (HYP) encapsulated on Pluronic® P123 (HYP/P123) against breast cancer cells (MCF-7) and non-tumorigenic breast cells (MCF-10A). METHODS: Cell internalization and subcellular distribution of HYP/P123 was confirmed by fluorescence microscopy. The phototoxicity and citototoxicity of HYP/P123 was assessed by trypan blue exclusion assay in the presence and absence of light. Long-term cytotoxicity was performed by clonogenic assay. Cell migration was determined by the wound-healing assay. Apoptosis and necrosis assays were performed by annexin VFITC/ propidium Iodide (PI) by fluorescence microscopy. RESULTS: Our results showed that HYP/P123 micelles had high stability and high rates of binding to cells, which resulted in the selective internalization in MCF-7, indicating their potential to permeate the membrane of these cells. Moreover, HYP/P123 micelles accumulated in mitochondria and endoplasmic reticulum organelles, resulting in the photodynamic cell death by necrosis. Additionally, HYP/P123 micelles showed effective and selective time- and dose dependent phototoxic effects on MCF-7 cells but little damage to MCF-10A cells. HYP/P123 micelles inhibited the generation of cellular colonies, indicating a possible capability to prevent the recurrence of breast cancer. We also demonstrated that HYP/P123 micelles inhibit the migration of tumor cells, possibly by decreasing their ability to form metastases. CONCLUSION: Taken together, the results presented here indicate a potentially useful role of HYP/P123 micelles as a platform for HYP delivery to more specifically and effectively treat human breast cancers through photodynamic therapy, suggesting they are worthy for in vivo preclinical evaluations.

Physiologically-Based Pharmacokinetic Model on the Oral Drug Absorption in Roux-en-Y Gastric Bypass Bariatric Patients: Amoxicillin Tablet and Suspension.

The potential of a physiologically-based pharmacokinetic (PBPK) model to predict oral amoxicillin bioavailability, by considering the physiological changes after "Roux-en-Y gastric bypass" (RYGB) surgery in bariatric patients, was evaluated. A middle-out approach for parameter estimations was undertaken using in vitro, in situ, and in vivo data. The observed versus predicted plasma concentrations and the model sensitivity of the simulated parameters of AUC0-inf and Cmax of amoxicillin (AMX) were used to confirm the reliability of the estimation. The model considers that a drug-transporter (Transp) in the initial segments of the normal intestine plays a significant role in the AMX absorption. A lower fraction absorbed (Fabs) was observed in RYGB patients (54.43% for suspension and 45.21% for tablets) compared to healthy subjects (77.48% capsule). Furthermore, the tablet formulation presented a lower dissolved fraction (Fd) and Fabs compared to the suspension formulation of AMX in RYGB patients (91.70% and 45.21% versus 99.92% and 54.43%, respectively). The AUC0-inf and Cmax were sensitive to changes in Rtintestine, PeffAMX, and Transp for both healthy and RYGB models. Additionally, AUC0-inf and Cmax were also sensitive to changes in the tlag parameter for tablet formulation in RYGB patients. The PBPK model showed a reduction in AMX bioavailability as a consequence of reduced intestinal length after RYGB surgery. Additionally, the difference in the predicted Fd and Fabs between suspension and tablet suggests that liquid formulations are preferable in postbariatric patients.

"Biotin-targeted mixed liposomes: A smart strategy for selective release of a photosensitizer agent in cancer cells".

The high incidence of cancer, necessity of treatment, and prognosis times are urgent issues that need to be addressed. In this work, we present DPPC liposomes coated with F127 triblock copolymers as a promising alternative in drug delivery systems for cancer therapy. The proposed mixed liposomes exhibit adequate size, high stability, and passive targeting that result from the EPR effect. An interesting strategy to obtain both passive and active targeting is the vectorization with a covalent bond between F127 and Biotin (a vitamin). Cancer cells can overexpress Biotin receptors, such as Avidin. Here, we evaluate the cytotoxic effects of the erythrosine-decyl ester (ERYDEC). This is a photosensitizer that can be utilized in photodynamic therapy (PDT) and incorporated in DPPC liposomes coated with F127 (F127/DPPC) and the biotinylated-F127 (F127-B/DPPC). The results showed that DPPC liposomes were efficiently mixed with common F127 or F127B, exhibiting adequate physical properties with simple and low-cost preparation. An HABA/Avidin assay showed the amount of Biotin available at the liposome surface. In addition, ERYDEC interaction with lipid vesicles showed high encapsulating efficiency and slow release kinetics. The ERYDEC monomeric species are represented by high light absorption and high singlet oxygen generation (1O2), which confirm the presence of the drug in its monomeric state, as required for PDT. The ERYDEC/liposome system showed high stability and absence of significant cytotoxic effects (absence of light) in fibroblasts of the Mus musculus cell line. In addition, phototoxicity studies showed that ERYDEC/liposomes were able to inhibit cancer cells. However, in the biotinylated system, the effect was much greater than the common F127 coating. This dramatically decreased the inhibitory concentration of CC50 and CC90. In addition, cellular uptake studies based on fluorescence properties of ERYDEC showed that a two-hour incubation period was enough for the uptake by the cell. Therefore, the new vectorized-coated liposome is a potential system for use in cancer treatments, considering that it is a theranostic platform.

Simultaneous liquid chromatography- tandem mass spectrometry method to quantify epicatechin and procyanidin B2 in rat plasma after oral administration of Trichilia catigua (catuaba) extract and its application to a pharmacokinetic study

Abstract Trichilia catigua A. Juss., Meliaceae, known as catuaba in Brazil, is traditionally used for the treatment of stress, sexual impotence and memory deficits. To our knowledge, there is no analytical method described in literature for simultaneous quantification of catuaba extract marker substances in biological matrices. The aim of this study was to develop and validate a bioanalytical method by LC-MS/MS to quantify epicatechin and procyanidin B2 in rat plasma after administration of standardized extract of T. catigua. Chromatographic separation was achieved with a C18 column, methanol and 0.1% aqueous formic acid at a flow rate of 0.25 ml/min. Detection was performed using electrospray ionization in negative mode. The lower limits of quantification were 5 ng/ml and 12.5 ng/ml for procyanidin B2 and epicatechin, respectively. Intra- and inter-day assays variability were less than 15%. The extraction recovery was 104% for epicatechin and 74% for procyanidin B2 using one-step liquid-liquid extraction with ethyl acetate. Epicatechin and PB2 were detected in plasma up to 300 min after oral administration of 400 mg/kg of standardized extract of T. catigua in rats. This rapid and sensitive method for the analysis of the epicatechin and procyanidin B2 in rat plasma can be applied to pharmacokinetic studies.

Reduced bioavailability of oral amoxicillin tablets compared to suspensions in Roux-en-Y gastric bypass bariatric subjects.

AIMS: To evaluate the relative bioavailability of oral amoxicillin (AMX) tablets in comparison to AMX suspension in Roux-en-Y gastric bypass bariatric subjects. METHODS: A randomized, double-blind, cross-over study was performed on the bioavailability of oral AMX tablets and suspension in Roux-en-Y gastric bypass subjects operated at least 3 months previously . Doses of 875 mg of the AMX tablet or 800 mg of the AMX suspension were given to all the subjects, allowing a washout of 7 days between the periods. Blood samples were collected at 0, 0.25, 0.5, 1, 1.5, 2, 4, 6 and 8 hours after drug administration and the AMX levels were quantified by liquid chromatography coupled with triple quadrupole tandem mass spectrometry. The pharmacokinetic parameters were calculated by noncompartmental analysis, normalized to an 875 mg dose and the bioavailability of the AMX from the tablets was compared to that from the suspension formulation. RESULTS: Twenty subjects aged 42.65 ± 7.21 years and with a body mass index of 29.88 ± 4.36 kg/m2 were enrolled in the study. The maximum AMX plasma concentration of the tablets and the suspension (normalized to 875 mg) were 7.42 ± 2.99 mg/L and 8.73 ± 3.26 mg/L (90% confidence interval of 70.71-99.11), and the total area under the curve from time zero to infinity were 23.10 ± 7.41 mg.h/L and 27.59 ± 8.32 mg.h/L (90% confidence interval of 71.25-97.32), respectively. CONCLUSION: The tablets presented a lower bioavailability than the suspension formulation and the total absorbed amount of AMX in these subjects was lower in comparison to the standard AMX absorption rates in nonbariatric subjects, regardless of the formulation.

Liposome and polymeric micelle-based delivery systems for chlorophylls: Photodamage effects on Staphylococcus aureus.

Chlorophyll derivatives (Chls), loaded in F-127 polymeric micelles and DPPC liposomes as drug delivery systems (DDS), have been shown to be remarkable photosensitizers for photodynamic inactivation (PDI). Assays of photoinactivation of Staphylococcus aureus bacteria (as biological models) showed that the effectiveness of Chls in these nanocarriers is dependent on photobleaching processes, photosensitizer locations in DDS, singlet oxygen quantum yields, and Chl uptake to bacteria. These are factors related to changes in Chl structure, such as the presence of metals, charge, and the phytyl chain. The photodynamic activity was significantly greater for Chls without the phytyl chain, i.e., phorbides derivatives. Furthermore, the inactivation of S. aureus was increased by the use of liposomes compared to micelles. Therefore, this research details and shows the high significance of the Chl structure and delivery system to enhance the photodynamic activity. It also highlights the chlorophylls (particularly phorbides) in liposomes as promising photosensitizers for PDI.

Thermal Magnetic Field Activated Propolis Release From Liquid Crystalline System Based on Magnetic Nanoparticles.

Intra-periodontal pocket drug delivery systems, such as liquid crystalline systems, are widely utilized improving the drug release control and the therapy. Propolis is used in the treatment of periodontal diseases, reducing the inflammatory and infectious conditions. Iron oxide magnetic nanoparticles (MNPs) can improve the treatment when an alternating external magnetic field (AEMF) is applied, increasing the local temperature. The aim of this study was to develop a liquid crystalline system containing MNPs for intra-periodontal pocket propolis release. MNPs were prepared using iron salts and the morphological, size, thermal, x-ray diffraction, magnetometry, and Mössbauer spectroscopy analyses were performed. Cytotoxicity studies using Artemia salina and fibroblasts were also accomplished. The systems were prepared using polyoxyethylene (10) oleyl ether, isopropyl myristate, purified water, and characterized by polarized optical microscopy, rheometry, and in vitro drug release profile using a periodontal pocket simulator apparatus. The antifungal activity of the systems was investigated against Candida spp. using an AEMF. MNPs displayed nanometric size, were monodisperse, and they displayed very low cytotoxicity. Microscopically homogeneous formulations were obtained displaying important physicochemical and biological properties. The system displayed prolonged release of propolis and important in vitro fungicide activity, which was increased when the AEMF was applied, indicating a potentially alternative therapy for the treatment of the periodontal disease.

Thalassemia major phenotype caused by HB Zürich-Albisrieden [α2 59(E8) Gly > Arg (HBA2:C.178G > C)] in a Brazilian child.

Hemoglobin (Hb) Zürich-Albisrieden (ZA) [α2 59(E8) Gly > Arg; HBA2:c.178G > C] is a rare and highly unstable α-chain variant. A few simple and compound heterozygotes (αZA α/αα and -/αZA α, respectively) have been described so far in Switzerland and China. We describe here a case of homozygosity for the Hb ZA mutation (αZA α/αZA α) in a Brazilian child with severe congenital hemolytic anemia and ineffective erythropoiesis.

Design and Characterization of Mucoadhesive Gelatin-Ethylcellulose Microparticles for the Delivery of Curcumin to the Bladder.

BACKGROUND: Bladder cancer is the second type of malignant carcinoma of the urinary tract. The treatment is time-consuming and requires maintenance doses of the drug for long period of time with important side effects. Curcumin has shown evident clinical advances in the treatment of cancer. The technology of microencapsulation and the use of mucoadhesive materials can contribute to modify the delivery and improve the bioavailability of curcumin. OBJECTIVE: The aim of this study was to design and characterize mucoadhesive microparticles containing curcumin using multivariate analysis and the spray-drying technique. METHODS: A factorial design 32+1 was employed to investigate the influence of gelatin, ethylcellulose, and curcumin on size, polydispersity index, drug content and entrapment efficiency. Microparticles were also evaluated by ATR-FTIR, X-ray diffraction, antioxidant activity, in-vitro release profile, exvivo mucoadhesion performance, and in-vitro cytotoxicity. RESULTS: Microparticles showed non-uniform surface, mean diameter from 2.73 µm to 4.62 µm and polydispersity index from 0.72 to 1.09, according to the different combinations of the independent factors. These independent variables also had a significant effect on the drug content. The highest values of drug trapping efficiency were obtained with the highest concentration of curcumin and polymers. Formulations displayed slow drug release and important antioxidant activity. The good mucoadhesive performance of microparticles was assessed by the falling film technique. Moreover, the formulations did not display in vitro toxicity against Artemia salina and Fibroblasts LM(TK). CONCLUSION: The design results were useful for developing of curcumin dosage form with good physicochemical characteristics and mucoadhesive properties for the bladder administration.

Rhesus θ-Defensin-1 Attenuates Endotoxin-induced Acute Lung Injury by Inhibiting Proinflammatory Cytokines and Neutrophil Recruitment.

Acute lung injury (ALI) is a clinical syndrome characterized by acute respiratory failure and is associated with substantial morbidity and mortality. Rhesus θ-defensin (RTD)-1 is an antimicrobial peptide with immunomodulatory activity. As airway inflammation and neutrophil recruitment and activation are hallmarks of ALI, we evaluated the therapeutic efficacy of RTD-1 in preclinical models of the disease. We investigated the effect of RTD-1 on neutrophil chemotaxis and macrophage-driven pulmonary inflammation with human peripheral neutrophils and LPS-stimulated murine alveolar macrophage (denoted MH-S) cells. Treatment and prophylactic single escalating doses were administered subcutaneously in a well-established murine model of direct endotoxin-induced ALI. We assessed lung injury by histopathology, pulmonary edema, inflammatory cell recruitment, and inflammatory cytokines/chemokines in the BAL fluid. In vitro studies demonstrated that RTD-1 suppressed CXCL8-induced neutrophil chemotaxis, TNF-mediated neutrophil-endothelial cell adhesion, and proinflammatory cytokine release in activated murine alveolar immortalized macrophages (MH-S) cells. Treatment with RTD-1 significantly inhibited in vivo LPS-induced ALI by reducing pulmonary edema and histopathological changes. Treatment was associated with dose- and time-dependent inhibition of proinflammatory cytokines (TNF, IL-1ß, and IL-6), peroxidase activity, and neutrophil recruitment into the airways. Antiinflammatory effects were demonstrated in animals receiving RTD-1 up to 12 hours after LPS challenge. Notably, subcutaneously administered RTD-1 demonstrates good peptide stability as demonstrated by the long in vivo half-life. Taken together, these studies demonstrate that RTD-1 is efficacious in an experimental model of ALI through inhibition of neutrophil chemotaxis and adhesion, and the attenuation of proinflammatory cytokines and gene expression from alveolar macrophages.

Rare α0-thalassemia deletions detected by MLPA in five unrelated Brazilian patients

Abstract Alpha-thalassemias are among the most common genetic diseases in the world. They are characterized by hypochromic and microcytic anemia and great clinical variability, ranging from a practically asymptomatic phenotype to severe anemia, which can lead to intrauterine or early neonatal death. Deletions affecting the α-globin genes, located on chromosome 16p13.3, are the main causes of α-thalassemia. Multiplex ligation-dependent probe amplification (MLPA) can be used to detect rearrangements that cause α-thalassemia, particularly large deletions involving the whole α cluster and/or deletions in the HS-40 region. Here, MLPA was used to investigate the molecular basis of α-thalassemia in five unrelated patients, three of whom had Hb H disease. In addition to the -α3.7 deletion identified in the patients with Hb H disease, four different α0 deletions removing 15 to 225 kb DNA segments were found: two of them remove both the α genes, one affects only the regulatory element (HS-40) region, and another one extends over the entire α cluster and the HS-40 region. These results illustrate the diversity of α-thalassemia deletions in the Brazilian population and highlight the importance of molecular investigation in cases that present with microcytosis and hypochromia without iron deficiency and normal or reduced Hb A2 levels..

Rare α0-thalassemia deletions detected by MLPA in five unrelated Brazilian patients.

Alpha-thalassemias are among the most common genetic diseases in the world. They are characterized by hypochromic and microcytic anemia and great clinical variability, ranging from a practically asymptomatic phenotype to severe anemia, which can lead to intrauterine or early neonatal death. Deletions affecting the α-globin genes, located on chromosome 16p13.3, are the main causes of α-thalassemia. Multiplex ligation-dependent probe amplification (MLPA) can be used to detect rearrangements that cause α-thalassemia, particularly large deletions involving the whole α cluster and/or deletions in the HS-40 region. Here, MLPA was used to investigate the molecular basis of α-thalassemia in five unrelated patients, three of whom had Hb H disease. In addition to the -α3.7 deletion identified in the patients with Hb H disease, four different α0 deletions removing 15 to 225 kb DNA segments were found: two of them remove both the α genes, one affects only the regulatory element (HS-40) region, and another one extends over the entire α cluster and the HS-40 region. These results illustrate the diversity of α-thalassemia deletions in the Brazilian population and highlight the importance of molecular investigation in cases that present with microcytosis and hypochromia without iron deficiency and normal or reduced Hb A2 levels..

Coinheritance of Hb Bristol-Alesha [ß67(E11)Val→Met; HBB: c.202G>A] and the α212 Patchwork Allele in a Brazilian Child with Severe Congenital Hemolytic Anemia.

Hb Bristol-Alesha [HBB: c.202G>A; ß 67 Val>Met] is a rare structural variant of hemoglobin (Hb) resulting from a GTG>ATG substitution at codon 67 of the ß-globin gene that leads to the replacement of valine by methionine in the corresponding position of the ß-globin chain. The methionine residue is subsequently modified to aspartic acid [ß67(E11)Val-Met→Asp], possibly by autoxidation mechanisms. This substitution prevents normal non-polar binding of Val67 to the heme group, resulting in molecular instability and severe hemolysis. We identified Hb Bristol-Alesha (in the heterozygous state), as the cause of severe congenital hemolytic anemia in an 11-month-old girl of mixed (native Indian and European) ethnic origin from the Midwestern region of Brazil, whose parents were clinically and hematologically normal. The mutation on the ß-globin gene was found to have been coinherited with the α212 patchwork allele.

Efficacy of Rhesus Theta-Defensin-1 in Experimental Models of Pseudomonas aeruginosa Lung Infection and Inflammation.

Chronic airway infection and inflammation contribute to the progressive loss of lung function and shortened survival of patients with cystic fibrosis (CF). Rhesus theta defensin-1 (RTD-1) is a macrocyclic host defense peptide with antimicrobial and immunomodulatory activities. Combined with favorable preclinical safety and peptide stability data, RTD-1 warrants investigation to determine its therapeutic potential for treatment of CF lung disease. We sought to evaluate the therapeutic potential of RTD-1 for CF airway infection and inflammation using in vitro, ex vivo, and in vivo models. We evaluated RTD-1's effects on basal and Pseudomonas aeruginosa-induced inflammation in CF sputum leukocytes and CF bronchial epithelial cells. Peptide stability was evaluated by incubation with CF sputum. Airway pharmacokinetics, safety, and tolerance studies were performed in naive mice. Aerosolized RTD-1 treatment effects were assessed by analyzing lung bacterial burdens and airway inflammation using an established model of chronic P. aeruginosa endobronchial infection in CF (ΔF508) mice. RTD-1 directly reduces metalloprotease activity, as well as inflammatory cytokine secretion from CF airway leukocyte and bronchial epithelial cells. Intrapulmonary safety, tolerability, and stability data support the aerosol administration route. RTD-1 reduced the bacterial lung burden, airway neutrophils, and inflammatory cytokines in CF mice with chronic P. aeruginosa lung infection. Collectively, these studies support further development of RTD-1 for treatment of CF airway disease.