Effect of treatment with growth hormone on body composition and metabolic profile of short children born small for gestational age / Efeito do tratamento com hormônio de crescimento na composição corporal e no perfil metabólico de crianças nascidas pequenas para a idade gestacional com baixa estatura

ABSTRACT Objective: To assess the effect of recombinant growth hormone (rGH) on body composition and metabolic profile of prepubertal short children born small for gestational age (SGA) before and after 18 months of treatment. Methods: It is a clinical, non-randomized, and paired study. Children born SGA, with birth weight and/or length <-2 standard deviations (SD) for gestational age and sex, prepubertal, born at full term, of both genders, with the indication for treatment with rGH were included. The intervention was performed with biosynthetic rGH at doses ranging from 0.03 to 0.05 mg/kg/day, administered subcutaneously, once a day at bedtime. Total lean mass (LM) and total fat mass (FM) were carried out using dual-energy X-ray absorptiometry (DXA), and the metabolic profile was assessed for insulin, glycemia, IGF-1 levels and lipid profile. Results: Twelve patients (nine girls, 8.17±2.39 y) were evaluated; three patients dropped out of the study. There was an increase of LM adjusted for length (LMI) (p=0.008), LMI standard deviation score (SDS) adjusted for age and sex (p=0.007), and total LM (p<0.001). The percentage of body fat (BF%) and abdominal fat (AF) remained unaltered in relation to the beginning of treatment. Among the metabolic variables, blood glucose remained within normal levels, and there was a reduction in the number of participants with altered cholesterol (p=0.023). Conclusions: The effect of rGH treatment was higher on LM than in FM, with increased LM adjusted for length and standardized for age and sex. Glycemia remained within the normal limits, and there was a decreased number of children with total cholesterol above the recommended levels.

RESUMO Objetivo: Avaliar o efeito do hormônio de crescimento recombinante (rHC) na composição corporal e no perfil metabólico de crianças pré-púberes com baixa estatura, nascidas pequenas para a idade gestacional (PIG) antes e depois de 18 meses de tratamento. Métodos: Estudo clínico, não randomizado e pareado. Foram incluídas crianças nascidas PIG, com peso e/ou altura ao nascer <-2 desvios padrão (DP) para idade gestacional e sexo, pré-púberes, nascidas a termo, de ambos os sexos, com indicação de tratamento com rGH. A intervenção foi realizada com rGH biossintético com doses variando de 0,03 a 0,05 mg/kg/dia, administrado por via subcutânea, uma vez ao dia ao deitar-se. A massa magra total (LM) e a massa gorda total (MG) foram determinadas por meio de absorciometria de raios X de dupla energia (DXA), e o perfil metabólico foi avaliado com dosagens de insulina, glicemia, IGF-1 e perfil lipídico. Resultados: Doze pacientes (nove meninas, 8,17±2,39 anos) foram avaliados; três pacientes abandonaram o estudo. Houve aumento da LM ajustada para estatura (LMI) (p=0,008), LMI standard deviation scores (SDS) ajustada para idade e sexo (p=0,007) e LM total (p<0,001). O percentual de gordura corporal (GC%) e gordura abdominal (AF) permaneceu inalterado em relação ao início do tratamento. Entre as variáveis metabólicas, a glicemia manteve-se na normalidade, e houve redução do número de participantes com colesterol alterado (p=0,023). Conclusões: O efeito do tratamento com HCr foi maior na MM do que na MG, com o aumento da MM ajustada para altura e padronizada para idade e sexo. A glicemia permaneceu normal e houve redução do número de crianças com colesterol total acima do recomendado.

Establishment and validation of bioactivity measurement method for recombinant human midkine / 药学学报

To establish and optimize a method for the detection of recombinant human midkine (rhMK) activity and verify its methodology, cell counting kit-8 (cck-8) method was used to measure the proliferation activity of rat knee chondrocytes. The specificity, accuracy, precision, linearity and robustness of the method were also verified in this study. The established method was proven to have good specificity because the buffer of rhMK and recombinant human interleukin-1 receptor antagonist have no obvious active effect; the recoveries of the samples with relative activities of 50%, 75%, 100%, 125%, 150% were in the range of 80.0% to 124.0% by statistical analysis, the relative standard deviations (RSD) of relative potency were all within 20%, the linear correlation coefficient, R2 ≥ 0.98, suggesting that the accuracy, precision and linearity of the method were good; the robustness correlation coefficient, R2 ≥ 0.92 and the ratio of maximum to minimum of sigmoidal dose-response were no less than 1.5, indicating that robustness of the methods was good. In conclusion, a bioactivity measurement method for rhMK was established and fully validated in this study and it provides a reliable method for the bioactivity analysis of rhMK routine samples during the development. This study was approved by the Animal Care and Use Committee of Shanghai Model Organisms Center, Inc. (approval number: 2019-0008-06).

Obtención de la proteína verde fluorescente recombinante y su anticuerpo policlonal Igy

La Proteína Verde Fluorescente (Green Fluorescent Protein, GFP) es ampliamente utilizada en ensayos in vivo e in vitro. Se han generado múltiples variantes de esta proteína para diversificar sus características, como la GFP-enhancer (EGFP) que emite una señal de fluorescencia 35 veces mayor en comparación con la proteína silvestre, siendo implementada como proteína fusión en estudios de localización y estabilidad estructural, entre otros. La detección de esta proteína y sus variantes puede ser directa o indirecta, mediante el uso de anticuerpos anti-GFP. Aunque el uso de GFP es generalizado y de evidente utilidad en investigación y en docencia, los insumos para su estudio exhiben un alto costo dado que deben ser importados, constituyendo un recurso limitado en Colombia. El presente trabajo reporta la clonación y expresión de la proteína recombinante 6xHisEGFP, cuya purificación se completó a partir de la fracción soluble e insoluble del sistema heterólogo Escherichia coli mediante cromatografía de afinidad a metales inmovilizados y electroforesis preparativa, respectivamente. La proteína purificada se implementó como antígeno para la producción de anticuerpos policlonales aviares (IgY) contra la EGFP, los cuales se obtuvieron desde los huevos colectados y el suero de las sangrías de las gallinas inmunizadas. En este sentido, la estrategia metodológica planteada constituye un avance en el desarrollo de un sistema biotecnológico para la producción nacional de herramientas moleculares como los anticuerpos policlonales aviares a bajo costo.

Green Fluorescent Protein (GFP) is widely used in in vivo and in vitro assays. Multiple variants of this protein have been generated to diversify its characteristics, such as the enhancer GFP (EGFP) that emits a 35-fold higher fluorescence signal compared to the wild-type protein, being implemented as a fusion reporter in localization and structural stability studies, among others. Detection of this protein can be direct or indirect, fusing anti-GFP antibodies. Although the use of GFP is generalized and of evident utility in research and teaching, the molecular tools for its study exhibit a high cost since they must be imported, constituting a limited resource in Colombia. This work reports the cloning and expression of the recombinant protein 6xHisEGFP, which purification was completed from the soluble and insoluble fraction of the heterologous Escherichia coli system by immobilized metal affinity chromatography and preparative SDS-PAGE, respectively. The purified protein was implemented as an antigen to produce avian polyclonal antibodies (IgY) against EGFP, which were obtained from collected eggs and blood serum from immunized hens. In this sense, the proposed methodological strategy constitutes an advance in the development of a biotechnological system for the national production of molecular tools such as avian polyclonal antibodies at low-cost.

The Use of Escherichia coli for Recombinant Human Insulin Production

Insulin is the essential hormone produced by the pancreas. which is accountable for sanctioning glucose we acquire from our food sources to be deposited in our body cells. Without insulin, our bodies cannot control blood sugar levels, so insulin is a vital hormone for survival. A diabetic person either does not produce insulin or is resilient to it for a multiple reasons. Because of this, they need insulin injections to process glucose. It has become stress-free for patients around the world to acquire insulin with the production of recombinant human insulin produced by Escherichia coli . This short review will provide an overview of the steps engaged in constructing recombinant human insulin utilizing the K12 strain of E. coli along with the prominence of recombinant insulin and why E. coli is most commonly used for insulin production.

Osteometabolic profile and bone mass in the transition phase: ethnic differences in Brazilians treated with somatropin during childhood

Abstract Objective: The main growth hormone action is to promote linear growth increasing protein synthesis stimulation and osteoblastic activity. Peak bone mass extends from adolescence to 30 years of age. Several factors may influence this acquisition and prevent fracture risk in adulthood, such as genetic potential, GH, ethnicity, and lifestyle factors. This study aims to compare bone mass and osteometabolic profile of white and Afro-descendant Brazilian adolescents in the transition phase, who were treated with human recombinant growth hormone in childhood. Methods: The authors selected 38 adolescents from the Transition Outpatient Clinic of the University of São Paulo. Lumbar spine and total body bone mineral density (BMD) and bone mineral content (BMC), serum calcium, 25-OH-vitamin D and bone markers were analyzed at the rhGH withdrawal. Results: The mean age was 16.8 ± 1.6 years. There were 21 Afro-descendant and 17 whites. Thirty-four percent of the sample presented vitamin D insufficiency, 66% inadequate calcium intake and 44.7% physical inactivity. The Afro-descendants showed a lower lumbar spine and total body Z scores than those of the whites (p = 0.04 and p = 0.03, respectively), as well as their mean body weight (p = 0.03). There were no differences in the remaining osteometabolic parameters. Conclusion: As most adolescents had vitamin D insufficiency, low calcium intake, and physical inactivity, calcium, and cholecalciferol supplementation and lifestyle changes should be encouraged. The Brazilian Afro-descendant may be a vulnerable group for low bone mass, requiring

Successful outcomes of surgery procedures in patient with Glanzmann´s Thrombasthenia

We report a 35-year-old female patient with Glanzmann's thrombasthenia (GT) and severe anemia due to abnormal uterine bleeding secondary to uterine myomatosis. She required several admissions of red blood cells and platelet transfusions. An elective subtotal hysterectomy with salpingo-oophorectomy was proposed and recombinant factor VII was required. Surgical and postoperative outcomes were successful, without surgical complications, bleeding, or hemogram alterations. 4 years later, she required tooth extraction because of periodontal disease and pulp necrosis. In Peru, reports of GT patients requiring major and minor surgical procedures are lacking, given the low disease prevalence and the difficulties related to surgery. The report of these successful cases becomes relevant to continue improving GT management.

Presentamos el caso de una paciente de 35 años con trombastenia de Glanzmann (GT) y anemia severa por sangrado uterino anormal secundario a miomatosis uterina. Requirió varias admisiones de transfusiones de glóbulos rojos y plaquetas. Se propuso histerectomía subtotal electiva con salpingo-ooforectomía y se requirió factor VII recombinante. Los resultados quirúrgicos y postoperatorios fueron exitosos, sin complicaciones quirúrgicas, sangrado ni alteraciones del hemograma. 4 años después, requirió extracción dental por enfermedad periodontal y necrosis pulpar. En Perú faltan reportes de pacientes con GT que requieran procedimientos quirúrgicos mayores y menores, dada la baja prevalencia de la enfermedad y las dificultades relacionadas con la cirugía. El reporte de estos casos de éxito cobra relevancia para seguir mejorando la gestión de GT

Angioedema after rt-PA infusion led to airway emergency: a case report of rescue treatment with fresh frozen plasma

Abstract The authors report the case of a 71-year-old woman presented to the Emergency Department with acute ischemic stroke. She was treated with rt-PA and interventional endovascular revascularization and developed rapidly progressing angioedema that led to emergency intubation. The standard treatment was not very effective and the swelling improved after infusion of fresh frozen plasma. Angioedema after rt-PA infusion could be a life-threatening emergency that requires quick airway management by skilled professionals. As this condition is triggered by several factors, such as unregulated histamine and bradykinin production, the traditional treatment recommended by the guidelines may not be sufficient and the use of FFP can be considered as a safe and valuable aid.

Molecular characterization & recombination analysis of complete enterovirus-88 isolated from acute flaccid paralysis cases in India

Background & objectives: Focus on non-polio enteroviruses (NPEVs) causing acute flaccid paralysis (AFP) due to myelitis has increased with the containment of the poliovirus. Enterovirus-B88 (EV-B88) has been associated with the AFP cases in Bangladesh, Ghana, South Africa, Thailand and India. In India, EV-B88 infection was linked to AFP a decade ago; however, to date, no complete genome has been made available. In this study, the complete genome sequence of EV-B88 was identified and reported from two different States (Bihar and Uttar Pradesh) in India using the next-generation sequencing technique. Methods: Virus isolation was performed on the three AFP suspected cases as per the WHO-recommended protocol. Samples showing cytopathic effects in the human Rhabdocarcinoma were labelled as NPEVs. Next-generation sequencing was performed on these NPEVs to identify the aetiological agent. The contiguous sequences (contigs) generated were identified, and reference-based mapping was performed. Results: EV-B88 sequences retrieved in our study were found to be 83 per cent similar to the EV-B88 isolate from Bangladesh in 2001 (strain: BAN01-10398; Accession number: AY843306.1). Recombination analyses of these samples demonstrate recombination events with sequences from echovirus-18 and echovirus-30. Interpretation & conclusions: Recombination events in the EV-B serotypes are known, and this work reconfirms the same for EV-B88 isolates also. This study is a step in increasing the awareness about EV-B88 in India and emphasizes future studies to be conducted in the identification of other types of EV present in India.

Effects of recombinant human erythropoietin on cerebral blood flow in preterm infants: arterial spin labeling magnetic resonance imaging evaluation / 中国新生儿科杂志

Objective:To study the effects of recombinant human erythropoietin (rhEPO) on cerebral blood flow (CBF) in preterm infants using arterial spin labeling (ASL) magnetic resonance imaging (MRI).Methods:From September 2021 to June 2022, preterm infants (gestational age ≤32 weeks, birth weight ≤1 500 g) admitted to NICU of our hospital within 24 h after birth were randomly assigned into rhEPO group and control group for this prospective study. The rhEPO group was given rhEPO (500 IU/kg iv, once every other day for 2 weeks) within 72 h after birth plus symptomatic supportive treatment. The control group received same amount of normal saline injection. Both groups received brain MRI, diffusion-weighted imaging and ASL at adjusted gestational age of 35~37 weeks and CBF values of interested areas were measured.Results:A total of 85 infants were enrolled, including 40 in the rhEPO group and 45 in the control group. No significant differences existed in the incidences of periventricular-intraventricular hemorrhage, periventricular leukomalacia, focal white matter injury and extensive white matter injury between the two groups ( P>0.05). The CBF values [ml/(100 g·min)] of frontal cortex [left 15.1±3.9 vs. 17.9±3.1, right 15.9 (12.5, 17.8) vs. 18.1(16.1,20.2)], temporal cortex [left 15.8±4.3 vs. 18.6±3.8, right 16.3(13.2,19.4) vs. 18.1(15.7,19.7)], occipital cortex (left 15.8±6.1 vs. 18.8±3.3, right 16.8±5.5 vs. 19.3±4.8), basal ganglia (left 24.7±7.2 vs. 28.7±6.2, right 26.0±7.9 vs. 29.3±6.4) and thalamus (left 32.7±11.8 vs. 37.9±8.6, right 32.1±11.6 vs. 37.6±10.2) in the rhEPO group were significantly lower than the control group ( P<0.05). No significant differences existed of CBF value at the parietal cortex between the two groups ( P>0.05). Conclusions:Early application of rhEPO can reduce CBF in premature infants, which may be related to the neuro-protective effects of EPO.

Advances in the pathogenesis and treatment of short stature in Noonan syndrome / 国际儿科学杂志

Noonan syndrome(NS)is an inherited disease involving multiple systems.The main clinical manifestations include distinctive facial features, short stature, heart defects, developmental delay and chest deformity.Short stature, reported in up to 70% of NS patients, is one of the main reasons NS patients seek medical treatment.The pathogenesis is associated with the up-regulation of RAS-mitogen activated protein kinase(RAS-MAPK)signal pathway.Further study is needed for some further specific mechanisms.Recombinant human growth hormone(rhGH)therapy has been approved for NS patients with short stature and has achieved a good therapeutic effect.However, the knowledge of drug dosage, influencing factors, long-term efficacy and risk of rhGH treatment is still insufficient.This paper reviews the pathogenesis and treatment of short stature in NS, providing help for the treatment and management of the disease.

Procine recombinant NK-lysin inhibits hepatocellular carcinoma metastasis by downregulating FKBP3 and inhibiting oxidative phosphorylation and glycolysis: a proteomic analysis / 南方医科大学学报

OBJECTIVE@#To investigate the potential mechanisms that mediate the inhibitory effect of porcine recombinant NKlysin (prNK-lysin) against liver cancer cell metastasis.@*METHODS@#HPLC-tandem mass spectrometry was used to identify the differentially expressed proteins in prNK-lysin-treated hepatocellular carcinoma SMMOL/LC-7721 cells in comparison with the control and PBS-treated cells. GO functional annotation and KEGG pathway analysis of the differentially expressed proteins were performed using GO and KEGG databases. RT-qPCR was used to determine the mRNA expression levels of polypeptide-N-acetylgalactosaminotransferase 13 (GALNT13), transmembrane protein 51 (TMEM51) and FKBP prolyl isomerase 3 (FKBP3) in the cells, and the protein expression of FKBP3 was verified using Western blotting.@*RESULTS@#Proteomic analysis identified 1989 differentially expressed proteins in prNK-lysin-treated cells compared with the control cells, and 2753 compared with PBS-treated cells. Fifteen proteins were differentially expressed between PBS-treated and the control cells, and 1909 were differentially expressed in prNK- lysin group compared with both PBS and control groups. These differentially expressed proteins were involved mainly in the viral process, translational initiation and RNA binding and were enriched mainly in ribosome, protein process in endoplasmic reticulum, and RNA transport pathways. RT-qPCR showed that compared with the control group, prNK-lysin treatment significantly increased the mRNA expressions of GALNT13 (P < 0.05) and TMEM51 (P < 0.01) and lowered FKBP3 mRNA expression (P < 0.05). Western blotting also showed a significantly decreased expression of FKBP3 protein in prNK-lysin-treated cells (P < 0.001).@*CONCLUSION@#Treatment with prNK-lysin causes significant changes in protein expression profile of SMMOL/LC-7721 cells and inhibits hepatocellular carcinoma metastasis by downregulating FKBP3 protein and affecting the cellular oxidative phosphorylation and glycolysis pathways.

Comparison of clinical efficacy of intravitreal injection of Conbercept and Aflibercept in the treatment of wet age-related macular degeneration / 国际眼科杂志(Guoji Yanke Zazhi)

AIM: To compare the clinical efficacy of conbercept and aflibercept in the treatment of wet age-related macular degeneration(wARMD)based on 4 consecutive intravitreal injections.METHODS: The clinical data of 108 patients(108 eyes)who were diagnosed as wARMD and treated with intravitreal injection at our hospital from January 2019 to January 2021 were retrospectively analyzed. They were divided into conbercept group(54 cases, 54 eyes)and aflibercept group(54 cases, 54 eyes)according to the different injectable drugs. All patients received intravitreal injection once a month, with four consecutive injections. Follow up for 12mo to observe best corrected visual acuity(BCVA), central macular thickness(CMT), complications and recurrence before and after injection.RESULTS: BCVA and CMT of patients in the two groups at 1, 2, 5 and 8mo after injection had no between-group differences(P&#x0026;#x003E;0.05), but both were significantly improved compared with those before injection(P&#x0026;#x003C;0.05). By the end of follow-up, conjunctival hemorrhage occurred in 2 eyes of the conbercept group at the early stage, and increased intraocular pressure and conjunctival hemorrhage occurred respectively in 2 eyes of the aflibercept group. There were no serious complications related to drug injection such as retinal detachment, complicated cataract, endophthalmitis and retinal pigment epithelial tear in the two groups, and there was no difference in the recurrence rate between the two groups(7% vs. 6%, P=1.000).CONCLUSION: On the basis of continuous 4 times of intravitreal injection, both conbercept and aflibercept are safe and effective in the treatment of wARMD, and the efficacy is even.

A novel bioassay to assess bioactivity of PEG-rhGH using Nb2-11 cells / 药学学报

In this study, we established a novel bioassay to determine the activity of polyethylene glycolated recombinant human growth hormone (PEG-rhGH) using Nb2-11 cells. We performed experimental condition optimization and methodological verification, and then detected the relative potency of PEG-rhGH products using this method. We demonstrated that the bioactivity of PEG-rhGH in promoting Nb2-11 cell proliferation displays a dose-response relationship, which conformed to the four-parameter model. Using PEG-rhGH reference as a control, we analyzed the relative potency of six batches of PEG-rhGH products, as well as linearity, regression and parallelism of the obtained curves. The relative potency of six batches of PEG-rhGH products was 95% to 105%. These results implied that the new bioassay established may be employed in quality control of PEG-rhGH products.

Collaborative study to evaluate a reporter gene assay for recombinant human follicle-stimulating hormone bioactivity / 药学学报

The goal of this work was to explore the prospect of standardized application of an in-vitro bioactivity assay for recombinant human follicle-stimulating hormone based on a reporter gene. The relative accuracy, intermediate precision, linearity and applicable range of the method were validated according to the General Rules of Chinese Pharmacopoeia 2020 edition Volume IV (9401). Three laboratories used this method to determine the in-vitro biological activities of six batches of drug product and three batches of drug substance manufactured by two different companies. The consistency of the potency determined by three laboratories, the intra-laboratory precision and inter-laboratory precision were analyzed. The method was optimized during the collaborative validation. The results of method validation meet the requirements of the General Rules of Chinese Pharmacopoeia 2020 edition Volume IV (9401). Aiming to resolve the problems found in the collaborative validation, the medium for cell seeding, the pre-diluted buffer solution of standard and sample, and the means of removing and discarding supernatant after stimulation were optimized. After optimization, there was no significant difference in the bioactivity among the different laboratories (P > 0.05), indicating statistical equivalency. Intra-laboratory and inter-laboratory precision were good and the geometric coefficient of variation (GCV%) were both less than 15%. In conclusion, the reporter gene assay has good intra-laboratory repeatability and inter-laboratory reproducibility and is suitable for analyzing recombinant human follicle-stimulating hormone drug product and drug substance by different manufacturers. It is expected to be used as a standardized method for the determination of the in-vitro bioactivity of such products.

Clinical comprehensive evaluation of recombinant Mycobacterium tuberculosis fusion protein / 中国药房

OBJECTIVE To evaluate the effectiveness， safety， economy， innovation， suitability and accessibility of recombinant Mycobacterium tuberculosis fusion protein （EC）， and to provide evidence for selecting skin detection methods for tuberculosis infection diagnosis and auxiliary diagnosis of tuberculosis. METHODS The effectiveness and safety of EC compared with purified protein derivative of tuberculin （TB-PPD） were analyzed by the method of systematic review. Cost minimization analysis， cost-effectiveness analysis and cost-utility analysis were used to evaluate the short-term economy of EC compared with TB-PPD， and cost-utility analysis was used to evaluate the long-term economy. The evaluation dimensions of innovation， suitability and accessibility were determined by systematic review and improved Delphi expert consultation， and the comprehensive score of EC and TB-PPD in each dimension were calculated by the weight of each indicator. RESULTS The scores of effectiveness， safety， economy， innovation and suitability of EC were all higher than those of TB-PPD. The affordability scores of the two drugs were consistent， while the availability score of EC was lower than those of TB-PPD. After considering dimensions and index weight， the scores of effectiveness， safety， economy， innovation， suitability， accessibility and the comprehensive score of EC were all higher than those of TB-PPD. CONCLUSIONS Compared with TB-PPD， EC performs better in all dimensions of effectiveness， safety， economy， innovation， suitability and accessibility. However， it is worth noting that EC should further improve its availability in the dimension of accessibility.

Effect of Recombinant Human Thrombopoietin (rhTPO) on Long-term Hematopoietic Recovery in Mice with Acute Radiation Sickness and Relative Mechanism / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect and relative mechanism of Recombinant Human Thrombopoietin (rhTPO) on long-term hematopoietic recovery in mice with acute radiation sickness.@*METHODS@#Mice were intramuscularly injected with rhTPO (100 μg/kg) 2 hours after total body irradiation with 60Co γ-rays (6.5 Gy). Moreover, six months after irradiation, peripheral blood, hematopoietic stem cells (HSC) ratio, competitive transplantation survival rate and chimerization rate, senescence rate of c-kit+ HSC, and p16 and p38 mRNA expression of c-kit+ HSC were detected.@*RESULTS@#Six months after 6.5 Gy γ-ray irradiation, there were no differences in peripheral blood white blood cells, red blood cells, platelets, neutrophils and bone marrow nucleated cells in normal group, irradiated group and rhTPO group (P>0.05). The proportion of hematopoietic stem cells and multipotent progenitor cells in mice of irradiated group was significantly decreased after irradiation (P<0.05), but there was no significant changes in rhTPO group (P>0.05). The counts of CFU-MK and BFU-E in irradiated group were significantly lower than that in normal group, and rhTPO group was higher than that of the irradiated group(P<0.05). The 70 day survival rate of recipient mice in normal group and rhTPO group was 100%, and all mice died in irradiation group. The senescence positive rates of c-kit+ HSC in normal group, irradiation group and rhTPO group were 6.11%, 9.54% and 6.01%, respectively (P<0.01). Compared with the normal group, the p16 and p38 mRNA expression of c-kit+ HSC in the irradiated mice were significantly increased (P<0.01), and it was markedly decreased after rhTPO administration (P<0.01).@*CONCLUSION@#The hematopoietic function of mice is still decreased 6 months after 6.5 Gy γ-ray irradiation, suggesting that there may be long-term damage. High-dose administration of rhTPO in the treatment of acute radiation sickness can reduce the senescence of HSC through p38-p16 pathway and improve the long-term damage of hematopoietic function in mice with acute radiation sickness.

Effect of Recombinant Human Thrombopoietin (rhTPO) on Hematopoietic Reconstitution in Allogeneic Hematopoietic Stem Cell Transplantation Model / 中国实验血液学杂志

OBJECTIVE@#To explore the effect of recombinant human thrombopoietin (rhTPO) on hematopoietic reconstruction in allogeneic hematopoietic stem cell transplantation (allo-HSCT) model.@*METHODS@#The C57BL/6 mice were employed as the donors, and BALB/c mice as recipients. The bone marrow mononuclear cells of the donor mice were extracted and pretreated, which then were injected with 5×106 per mouse through the tail vein of the recipient to establish an allo-HSCT model. The implantation of hematopoietic stem cells in the recipient mice was detected by flow cytometry on the 28th day after transplantation. Next, the successfully modeled recipient mice were randomly divided into experimental group and control group. The rhTPO was injected into mice in the experimental group on the first day after transplantation, while the saline was injected into mice in the control group. Both groups were injected for 14 consecutive days. The peripheral blood and bone marrow hematopoiesis of the two groups were observed on day 1, 3, 7, 14, and 21 after transplantation.@*RESULTS@#The expression rate of H-2Kb in the bone marrow of recipient mice was 43.85% (>20%) on the 28th day after transplantation, which indicated that the recipient mice were successfully chimerized. Meanwhile, counts of PLTs on the day 3, 7, 14, and 21 after transplantation in the experimental group were higher than those in the control group with statistical significances (P<0.05). In addition, hematopoietic function of bone marrow was suppressed in both groups on day 1, 3 and 7 after transplantation, but hematopoietic bone marrow hyperplasia was better in the experimental group than in the control group. On day 14 and 21 after transplantation, the hematopoietic function of bone marrow in the two groups was recovered, and the experimental group showed more obvious than the control group.@*CONCLUSION@#rhTPO can effectively stimulate the production of PLTs and facilitate the recovery of white blood cells and hemoglobin after allo-HSCT, and promote hematopoietic recovery and reconstitution of bone marrow.

Seleção de antígenos de Leishmania infantum por imunoproteômica para emprego no diagnóstico da leishmaniose visceral humana / Selection of Leishmania infantum antigens by immunoproteomics for use in the diagnosis of human visceral leishmaniasis

O controle da leishmaniose visceral (LV) requer um diagnóstico e tratamento adequados, uma vez que o diagnóstico preciso é fundamental para um regime medicamentoso eficaz para os pacientes. Nesse contexto, as ferramentas biotecnológicas devem ser aprimoradas para o manejo clínico e a avaliação epidemiológica da doença. No entanto, existem limitações relacionadas com a sensibilidade e/ou especificidade dos antígenos usados atualmente, mostrando a necessidade de identificação de novas moléculas para serem testadas em um diagnóstico sorológico mais sensível e específico. Neste sentido, no presente estudo, uma abordagem imunoproteômica foi usada para identificar proteínas antigênicas das formas promastigotas e amastigotas da espécie Leishmania infantum, causadora de LV em nosso país, por meio de seu reconhecimento por anticorpos em soros de pacientes com a doença. Amostras de indivíduos saudáveis residentes em região endêmica da doença e de pacientes com Doença de Chagas foram utilizadas com a função de se obter proteínas mais específicas ao parasito Leishmania para serem avaliadas no diagnóstico da LV. Como resultados obtidos, um total de 29 e 21 proteínas foram identificadas nos extratos de formas promastigotas e amastigotas dos parasitos, respectivamente. Para a validação da capacidade diagnóstica, duas proteínas, endonuclease III e GTP-binding protein, foram selecionadas, clonadas, expressas e purificadas para serem testadas em experimentos de ELISA. Os resultados dos testes mostraram valores de sensibilidade e especificidade superiores a 99,0% para a identificação da LV. Os antígenos ainda exibiram um diferencial ao apresentarem baixa reatividade sorológica em pacientes curados e tratados, sugerindo a possibilidade de que as mesmas possam ser aplicadas como marcadores prognósticos da doença. Em conclusão, o estudo imunoproteômico se mostrou eficaz na seleção de proteínas antigênicas de L. infantum e duas delas, endonuclease III e GTP-binding protein, foram bem avaliadas para o diagnóstico da LV frente a um painel sorológico, além de demonstrarem um potencial para monitoramento de pacientes com LV após o tratamento.

The control of visceral leishmaniasis (VL) requires an adequate diagnosis and treatment, since an accurate diagnosis is essential for an effective medication regimen for patients. In this context, biotechnological tools must be improved for the clinical management and epidemiological assessment of the disease. However, there are limitations related to the sensitivity and / or specificity of the antigens currently used, showing the necessity to identify new molecules to be tested in a more sensitive and specific serological diagnosis. In this sense, in the present study, an immunoproteomics approach was used to identify antigenic proteins of the Leishmania infantum promastigote and amastigote forms, which causes VL in our country, through its recognition by antibodies in sera of patients with the disease. Samples from healthy individuals living in an endemic region of the disease and from patients with Chagas disease were used to obtain more specific proteins for the Leishmania parasite, aiming their future application in the VL diagnosis. As results obtained, a total of 29 and 21 proteins were identified in the extracts of parasitic promastigotes and amastigotes, respectively. For validation of the diagnostic capacity, two proteins, endonuclease III and GTP-binding protein, were selected, cloned, expressed and purified to be tested in ELISA experiments. The test results showed sensitivity and specificity values greater than 99.0% for the identification of VL. The antigens also exhibited a differential when presenting low serological reactivity in cured and treated patients, suggesting the possibility that they can be applied as prognostic markers of the disease. In conclusion, the immunoproteomic study proved to be effective in the selection of L. infantum antigenic proteins and two of them, endonuclease III and GTP-binding protein, were well evaluated for the diagnosis of VL against a serological panel, in addition, demonstrating a potential for monitoring patients with VL after treatment.

Towards better antivenoms: navigating the road to new types of snakebite envenoming therapies

Abstract Snakebite envenoming is a significant global health challenge, and for over a century, traditional plasma-derived antivenoms from hyperimmunized animals have been the primary treatment against this infliction. However, these antivenoms have several inherent limitations, including the risk of causing adverse reactions when administered to patients, batch-to-batch variation, and high production costs. To address these issues and improve treatment outcomes, the development of new types of antivenoms is crucial. During this development, key aspects such as improved clinical efficacy, enhanced safety profiles, and greater affordability should be in focus. To achieve these goals, modern biotechnological methods can be applied to the discovery and development of therapeutic agents that can neutralize medically important toxins from multiple snake species. This review highlights some of these agents, including monoclonal antibodies, nanobodies, and selected small molecules, that can achieve broad toxin neutralization, have favorable safety profiles, and can be produced on a large scale with standardized manufacturing processes. Considering the inherent strengths and limitations related to the pharmacokinetics of these different agents, a combination of them might be beneficial in the development of new types of antivenom products with improved therapeutic properties. While the implementation of new therapies requires time, it is foreseeable that the application of biotechnological advancements represents a promising trajectory toward the development of improved therapies for snakebite envenoming. As research and development continue to advance, these new products could emerge as the mainstay treatment in the future.

Ensayo controlado y aleatorizado del uso de Heberkinasa® en el derrame pleural paraneumónico en niños / Randomized Controlled Trial of the Use of Heberkinase® in Parapneumonic Pleural Effusion in Children

Introducción: El derrame pleural paraneumónico resulta la complicación más frecuente de la neumonía bacteriana, de manejo complejo y muchas veces quirúrgico. No existen publicaciones en Cuba provenientes de ensayos clínicos controlados y aleatorizados ni del uso de la estreptoquinasa recombinante (Heberkinasa®) en el derrame pleural. Objetivo: Evaluar la eficacia y la seguridad de la Heberkinasa® en el tratamiento del derrame pleural paraneumónico complicado complejo y el empiema en niños. Métodos: Ensayo clínico fase III, abierto, aleatorizado (2:1), en grupos paralelos y controlado. Se concluyó la inclusión prevista de 48 niños (1-18 años de edad), que cumplieron los criterios de selección. Los progenitores otorgaron el consentimiento informado. Los pacientes se distribuyeron en dos grupos: I- experimental: terapia estándar y administración intrapleural diaria de 200 000 UI de Heberkinasa® durante 3-5 días y II-control: tratamiento estándar. Las variables principales: necesidad de cirugía y la estadía hospitalaria. Se evaluaron los eventos adversos. Resultados: Ningún paciente del grupo I-experimental requirió cirugía, a diferencia del grupo II-control en el que 37,5 por ciento necesitó cirugía video-toracoscópica, con diferencia altamente significativa. Se redujo la estadía hospitalaria (en cuatro días), las complicaciones intratorácicas y las infecciones asociadas a la asistencia sanitaria en el grupo que recibió Heberkinasa®. No se presentaron eventos adversos graves atribuibles al producto. Conclusiones: La Heberkinasa® en el derrame pleural paraneumónico complicado complejo y empiema resultó eficaz y segura para la evacuación del foco séptico, con reducción de la necesidad de tratamiento quirúrgico, de la estadía hospitalaria y de las complicaciones, sin eventos adversos relacionados con su administración(AU)

Introduction: Paraneumonic pleural effusion is the most frequent complication of bacterial pneumonia, with complex and often surgical management. There are no publications in Cuba from randomized controlled clinical trials or the use of recombinant streptokinase (Heberkinase®) in pleural effusion. Objective: To evaluate the efficacy and safety of Heberkinase® in the treatment of complex complicated parapneumonic pleural effusion and empyema in children. Methods: Phase III, open-label, randomized (2:1), parallel-group, controlled clinical trial. The planned inclusion of 48 children (1-18 years of age), who met the selection criteria, was completed. Parents gave informed consent. The patients were divided into two groups: I-experimental: standard therapy and daily intrapleural administration of 200,000 IU of Heberkinase® for 3-5 days; and II-control: standard treatment. The main variables: need for surgery and hospital stay. Adverse events were evaluated. Results: No patient in group I-experimental required surgery, unlike group II-control in which 37.5 percent required video-assisted thoracoscopic surgery, with a highly significant difference. Hospital stay (to 4 days), intrathoracic complications and infections associated to healthcare in the group that received Heberkinase® was reduced. No serious adverse events attributable to the product occurred. Conclusions: Heberkinase® in complex complicated parapneumonic pleural effusion and empyema was effective and safe for the draining of the septic focus, with reduction of the need for surgical treatment, hospital stay and complications, with no adverse events related to its administration(AU)