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This manuscript is intended to provide a best practice approach to accurately and consistently assess toxicant-induced bone marrow effects of test articles. In nonclinical toxicity studies, complete blood count data in conjunction with the histological examination of the bone marrow are recommended as the foundation for assessing the effect of test...
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... BM smears were prepared using the push-slide technique [42]. In brief, one drop of BM was placed at one extremity of a glass slide and was then spread in an even film using a second glass slide (held at an angle) as a spreader. ...
... Slides were then rinsed 3 times with pH 6.8 water before being air-dried. A qualitative and quantitative cytological evaluation of all the major lineages was performed by an experienced clinical pathologist on 300 to 500 cells [42]. ...
... Except for some BM samples, the percentages of the major BM lineages were indeed within reported ranges for C57BL/6J mice [56] (Figure 8). Albeit the performed evaluation of BM was partial and not fully differential, by grouping several developmental stages, it is considered adequate to characterize the major cytological findings in the BM [42]. Furthermore, Svensson et al. showed no acute or long term (sacrifice at 6 months p.i.) BM toxicity in nude mice treated with higher activities of 177 Lu-DOTATATE (90 MBq), as measured by leucocyte count [57]. ...
Simple Summary
Peptide receptor radionuclide therapy with ¹⁷⁷Lu-DOTATATE is an efficient treatment for patients suffering from metastasized neuroendocrine tumours. Nevertheless, suboptimal effects have been observed in the majority of patients. Hence, strategies to improve ¹⁷⁷Lu-DOTATATE efficacy are desirable. Lu-177 induces oxidative stress, eventually leading to tumour cell death. Inhibition of the antioxidant defence mechanisms, using buthionine sulfoximine (BSO), represents an attractive strategy to increase ¹⁷⁷Lu-DOTATATE efficacy. In cells and an animal model, the combination of ¹⁷⁷Lu-DOTATATE and BSO was more effective than ¹⁷⁷Lu-DOTATATE alone. In addition, it did not result in additional toxicity. Targeting the antioxidant defence system opens new safe treatment combination opportunities with ¹⁷⁷Lu-DOTATATE.
Abstract
Peptide receptor radionuclide therapy with ¹⁷⁷Lu-DOTATATE improves the outcome of patients with somatostatin receptor (SSTR)-expressing neuroendocrine tumours. Nevertheless, stable disease has been the main response pattern observed, with some rare complete responses. Lu-177 exerts about two-thirds of its biological effects via the indirect effects of ionizing radiation that generate reactive oxygen species, eventually leading to oxidative damage and cell death. This provides a rationale for targeting the antioxidant defence system in combination with ¹⁷⁷Lu-DOTATATE. In the present study, the radiosensitizing potential and the safety of depleting glutathione (GSH) levels using buthionine sulfoximine (BSO) during ¹⁷⁷Lu-DOTATATE therapy were assessed in vitro and in vivo using a xenograft mouse model. In vitro, the combination resulted in a synergistic effect in cell lines exhibiting a BSO-mediated GSH decrease. In vivo, BSO neither influenced ¹⁷⁷Lu-DOTATATE biodistribution nor induced liver, kidney or bone marrow toxicity. In terms of efficacy, the combination resulted in reduced tumour growth and metabolic activity. Our results showed that disturbing the cell redox balance using a GSH synthesis inhibitor increased ¹⁷⁷Lu-DOTATATE efficacy without additional toxicity. Targeting the antioxidant defence system opens new safe treatment combination opportunities with ¹⁷⁷Lu-DOTATATE.
... Then after making a bone marrow smear stained by Giemsa (23). Also count M: E ratio and differential erythroid and myeloid series (24). Collected blood samples from the jugular vein, and kept them in an EDTA tube to estimate blood parameters (25). ...
... Moreover, metarubricyte had a small clumped nucleus because of a reduction in its size and deep red cytoplasm that were also described (6,34). The myeloid series description was fainter than Rubriblast in staining (24) and included; Myeloblast, myelocyte had granulated cytoplasm, metamyelocyte with a reduced nucleus, and band cell with characters of the horseshoe nucleus, and that was like previous observations (1). We showed the differentiated rubricate and myelocyte in a high percentage, and such results were reported because several mitotic divisions included more maturation until they passed to the following progenitor stage (35,36). ...
... We showed the differentiated rubricate and myelocyte in a high percentage, and such results were reported because several mitotic divisions included more maturation until they passed to the following progenitor stage (35,36). We did not include the other advanced maturation stages for the myelocytic and erythrocytic series to avoid mistakes with peripheral blood elements that may enter the bone marrow smear during the sample collection procedure (24,37). The peripheral blood is mixed with bone marrow specimens at the collecting sample. ...
The state of the peripheral blood reflects on bone marrow activity. Therefore, we need to know the normal red bone marrow, and it can be a reference for those interested in hematology. Calves of apparently healthy local breeds were used to examine the state of the bone marrow, take a sample of peripheral blood before slaughter, and then a bone marrow biopsy that was immediately after slaughter. The calves that showed typical values for the parameters of red blood cells and total and differential white blood cells were included in the study. The result of bone marrow smears for nine normal calves appeared to have a faint coloration of granulated myelocytic than erythroid cells. As well as the myeloid was less than the erythroid, and that M: E ratio was 0.91±0.03. The erythroid series, Rubriblast, and Rubricyte were present as the most common progenitor red cell, including basophilic, polychromic, and normochromic cytoplasm, metarubricyte as a reduced nucleus. The myeloid series: Myeloblast, myelocyte was the most common progenitor white cells with granulation as basophilic, eosinophilic and neutrophilic within their cytoplasm, metamyelocyte with shrunken nuclei and band cell with horseshoe nucleus. They rarely found the megakaryocyte, enormous multinucleate cell.
... Furthermore, the effect of potassium bromate was not pronounced in the creatine metabolism based on the data provided in this present study. The hematopoietic system is an essential system to assess toxicity in humans and animals [63,64]. This present study has demonstrated the haematological cells or haemopoietin apparatus were not prone to oxidative damage by potassium bromate. ...
Some therapeutic and beneficial health properties of the Theobroma cacao leaf have been documented. This study evaluated the ameliorative effect of Theobroma cacao-fortified feed against potassium bromate-induced oxidative damage in male Wistar rats. Thirty rats were randomly grouped into A-E. Except for E (the negative control), the rats in the other groups were administered 0.5 ml of 10 mg/kg body weight of potassium bromate daily using oral gavage and then allowed access to feed and water ad libitum. Groups B, C, and D were fed with 10 %, 20 %, and 30 % leaf-fortified feed respectively, while the negative and positive control (A) was fed with commercial feed. The treatment was carried out consecutively for fourteen days. In the liver and kidney, there was a significant increase (p
... When the primary and secondary haematologic dysfunction or abnormalities cannot be interpreted by peripheral blood smear investigation, the bone marrow assessment is useful and advised [5]. Moreover, the BM appraisal is recommended for studying certain malignancies or investigating abnormalities in the haematopoietic system [6]. ...
Haematopoiesis is very complicated process of formation, development, and differentiation of bone marrow cells and its evaluation is very valuable diagnostic tool for the veterinary practitioner. So, the aim of study is to evaluate the healthy bone marrow cytology during first three months old. Ninety pups were involved throughout the study according to study design. Growing pups were kept with their moms till the weaning day, then divided into three main groups (1, 2, and 3 months old). After reaching the appropriate study age, bone marrow samples were obtained, and bone marrow smears were carried out and stained with MGGS. The results revealed presence of all types of haematopoietic cells and there are increase in different erythroid progenitor and myeloid progenitor with the age progress. Furthermore, the smears showed different cell line production with different differentiation stages of both erythroid and myeloid cells. In conclusion the bone marrow cells differentiation and proliferation are related to age progress independently. Haematopoietic cells differentiation and proliferation are regulated by bone marrow microenvironment. Myeloid cells are directly related to age progress.
... IL-6 can either directly suppress erythropoiesis or inhibit erythropoiesis by interfering with iron homeostasis, the latter effect being well studied in models of anemia of inflammation (AI). During inflammation, IL-6 induces the hepatic expression of hepcidin 3 , the master regulator of iron metabolism 4,5 . Under physiological conditions, hepcidin expression is induced by high iron levels via the BMP/HJV/SMAD pathway 6,7 . ...
Anemia of cancer (AoC) with its multifactorial etiology and complex pathology is a poor prognostic indicator for cancer patients. One of the main causes of AoC is cancer-associated inflammation that activates mechanisms, commonly observed in anemia of inflammation, where functional iron deficiency and iron-restricted erythropoiesis is induced by increased hepcidin levels in response to IL-6 elevation. So far only a few AoC mouse models have been described, and most of them did not fully recapitulate the interplay of anemia, increased hepcidin levels and functional iron deficiency in human patients. To test if the selection and the complexity of AoC mouse models dictates the pathology or if AoC in mice per se develops independently of iron deficiency, we characterized AoC in Trp53floxWapCre mice that spontaneously develop breast cancer. These mice developed AoC associated with high IL-6 levels and iron deficiency. However, hepcidin levels were not increased and hypoferremia coincided with anemia rather than causing it. Instead, an early shift in the commitment of common myeloid progenitors from the erythroid to the myeloid lineage resulted in increased myelopoiesis and in the excessive production of neutrophils that accumulate in necrotic tumor regions. This process could neither be prevented by iron nor erythropoietin (EPO) treatment. Trp53floxWapCre mice are the first mouse model where EPO-resistant anemia is described and may serve as a disease model to test therapeutic approaches for a subpopulation of human cancer patients with normal or corrected iron levels that do not respond to EPO.
... Finally, it should be noted that the interpretation of these panels can be species-specific. For instance, when ''Sternebrae, femur or vertebrae (including bone marrow)" is required, this can be sternum or femur with cartilage for rodents, while the femur may be less suited for non-rodents (due to the less uniform presence of active marrow and increased collection difficulty) [47]. ...
The success of the messenger RNA-based COVID-19 vaccines of Moderna and Pfizer/BioNTech marks the beginning of a new chapter in modern medicine. However, the rapid rise of mRNA therapeutics has resulted in a regulatory framework that is somewhat lagging. The current guidelines either do not apply, do not mention RNA therapeutics, or do not have widely accepted definitions. This review describes the guidelines for preclinical biodistribution studies of mRNA/siRNA therapeutics and highlights the relevant differences for mRNA vaccines. We also discuss the role of in vivo RNA imaging techniques and other assays to fulfill and/or complement the regulatory requirements. Specifically, quantitative whole-body autoradiography, microautoradiography, mass spectrometry-based assays, hybridization techniques (FISH, bDNA), PCR-based methods, in vivo fluorescence imaging, and in vivo bioluminescence imaging, are discussed. We conclude that this new and rapidly evolving class of medicines demands a multi-layered approach to fully understand its biodistribution and in vivo characteristics.
... The criteria for histopathological evaluation were based on the International Harmonization of Nomenclature and Diagnostic Criteria for Lesions in Rats and Mice Vaccine xxx (xxxx) xxx for the cardiovascular system [12]. Cytological assessments of the bone marrow were performed based on the recommendations of the American Society for Veterinary Clinical Pathology and the Society for Toxicologic Pathology working group [13]. ...
ChAdOx1 nCoV-19 adenoviral vector vaccine (ChAd) against coronavirus disease 2019 has been associated with vaccine-induced thrombosis and thrombocytopenia (VITT), especially in young women who have presented with unusual localized thrombosis after receiving the vaccine. The pathogenesis of VITT remains incompletely understood. We tried to provide new insights into mechanisms underlying this phenomenon in the model of arterial thrombosis electrically induced in the carotid artery of female rats. At 28 days post-vaccination, ChAd induced SARS-CoV-2-specific neutralizing antibody responses in all animals. The analysis of the blood vessel/thrombus area showed slight luminal narrowing of the carotid artery with extravasation of blood in vaccinated rats. These small changes were not accompanied by differences in thrombus weight and composition. The vaccinated animals presented a slight increase (by around 14–24%) in platelet aggregation. ChAd did not significantly affect blood coagulation, platelet counts, and their activation markers. Unaffected thrombus formation, the lack of thrombocytopenia and all the measured blood and hemostasis parameters that predominantly stayed unchanged, indicate that the ChAd does not increase the risk of arterial thrombosis development in female rats.
... The Society's commitment to facilitate good regulatory decisions led to the formation of the STP Regulatory Affairs Committee, which later became the Scientific and Regulatory Policy Committee (SRPC). This committee communicates the viewpoint of toxicologic pathologists on technical topics (eg, appropriate collection, processing, and analysis of pathology samples [30][31][32][33][34][35] or regulatory positions (eg, performance of core pathology tasks such as histopathology data collection 36,37 and peer review 38,39 ) to health authorities and international consortia (eg, the Organisation for Economic Cooperation and Development [OECD]). These communications may be initiated by the SRPC to address a perceived need or may be undertaken specifically to answer a call for public comment by the health authority. ...
The Society of Toxicologic Pathology (STP, https://www.toxpath.org /) was founded in North America in 1971 as a nonprofit scientific and educational association to promote the professional practice of pathology as applied to pharmaceutical and environmental safety assessment. In the ensuing 50 years, the STP has become a principal global leader in the field. Society membership has expanded to include toxicologic pathologists and allied scientists (eg, toxicologists, regulatory reviewers) from many nations. In addition to serving membership needs for professional development and networking, major STP outreach activities include production of articles and presentations designed to optimize toxicologic pathology procedures (“best practice” recommendations), communicate core principles of pathology evaluation and interpretation (“points to consider” and “opinion” pieces), and participation in international efforts to harmonize diagnostic nomenclature. The STP has evolved into an essential resource for academic, government, and industrial organizations that employ and educate toxicologic pathologists as well as use toxicologic pathology data across a range of applications from assessing product safety (therapies, foods, etc) to monitoring and maintaining environmental and occupational health. This article recapitulates the important milestones and accomplishments of the STP during its first 50 years.
... The morphology of MDS myeloid cells were observed through Wright-Giemsa stained bone marrow smears. It was evaluated subjectively by light microscopy at low power (10 × objectives) for overall quality and distribution, and then was analyzed at high power (100 × oil objectives) for differential count including PMMBM count which was positively correlating with CD14 detected by flow cytometry (data not shown), with all cells in each containing field counted to maintain representative ratios of cell types [19]. All BM morphology findings were interpreted by two experienced and qualified clinical pathologists. ...
Background
Myelodysplastic syndromes (MDS) is a group of heterogeneous myeloid clonal diseases originating from hematopoietic stem cells. Clinically, elevated mature monocyte in bone marrow is often observed, but its clinical value still remains unclear.
Methods
We retrospectively analyzed a cohort of 216 MDS patients to explore the prognostic value of the percentage of mature monocyte in bone marrow (PMMBM). All patients were divided into elevated PMMBM group and the normal group by 6% PMMBM as the cut-off value.
Results
Our results showed that PMMBM> 6% was associated with inferior overall survival (OS) ( P = 0.026) along with higher-risk IPSS-R ( P = 0.025) and higher frequency of IDH2 mutation ( P = 0.007). Multivariate analyses showed that besides older age (> 60 years) for OS, gender (male) for OS, lower neutrophil count (< 0.8 × 10 ⁹ /L) for OS, higher bone marrow blast percentage (> 5%) for OS and LFS, poorer karyotype for OS, elevated PMMBM was also an independent adverse prognostic factor for OS in MDS ( P < 0.0001) but not for LFS ( P = 0.736).
Conclusions
These findings indicate that increased PMMBM may assists Revised International Prognostic Scoring System (IPSS-R) to predict a poor outcome and provide a novel evaluation factor for MDS patients especially when their karyotype analyses fail.
... The morphology of MDS myeloid cells were observed through Wright-Giemsa tinct bone marrow smear. It was evaluated subjectively by light microscopy at low power (10×objectives) for overall quality and distribution, and then was analyzed at high power (100×oil objectives) for differential count including PMMBM count, with all cells in each containing eld counted to maintain representative ratios of cell types [20] . All BM morphology ndings were interpreted by two experienced and quali ed clinical pathologists. ...
Background: Myelodysplastic syndromes (MDS) is a group of heterogeneous
myeloid clonal diseases originating from hematopoietic stem cells. Clinically, elevated mature monocyte is often observed, but its clinical value still remains unclear.
Methods: We retrospectively analyzed a cohort of 235 MDS patients to explore the prognostic value of the percentage of mature monocyte in bone marrow (PMMBM). All patients were divided into elevated PMMBM group and the normal group by 6% PMMBM as the cut-off value.
Results: Our results showed that PMMBM>6% was associated with inferior overall survival (OS) (P=0.007) and leukemia-free survival (LFS) (P=0.016) along with higher Revised International Prognostic Scoring System (IPSS-R) score (P<0.0001) and higher frequency of IDH2 mutation (P=0.001). Multivariate analyses showed that besides older age (>60 years), lower hemoglobin level (<10 g/dl), higher bone marrow blast percentage (>5%), poorer karyotype, elevated PMMBM was also an independent adverse prognostic factor for OS in MDS (P=0.049).
Conclusions: These findings indicate that increased PMMBM accompanied with a higher IPSS-R score may predict a poor outcome and provide a novel evaluation factor for MDS patients especially when their karyotype analyses fail.
