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OBJECTIVE: This study was carried out to assess the diagnostic utility of Bone Marrow Biopsy and Bone Marrow Culture in patients with PUO. METHODOLOGY: This retrospective cross-sectional study was conducted at Fauji Foundation Hospital, Rawalpindi. The clinical, laboratory and radiological records from January 2012 - December 2021 were retrieved from our hospital management information system for this study. Data from 63 patients with PUO were analyzed according to Petersdorf criteria. RESULTS: Out of 63 patients with PUO, BMB was diagnostic in 25 (39.68%) patients. The leading cause of PUO was infectious disorders (mycobacterium tuberculosis, visceral leishmaniasis, malaria), followed by inflammatory disorders and hematological malignancies. In 38 (60.31%) patients, BMB could not detect any underlying pathology. BMC showed growth in only 03(4.76%) patients, and the diagnostic yield of BMC was meagre. CONCLUSION: BMB is essential in establishing a diagnosis in patients with PUO. Infectious diseases are the leading cause of PUO, and BMC should not be included as a routine investigation in the initial evaluation of a patient with PUO. [ABSTRACT FROM AUTHOR]

Innate monocytes can be trained or reprogrammed to adopt distinct memory states, such as low-grade inflammation and immune exhaustion, bearing fundamental relevance to the pathogenesis of both acute diseases such as sepsis as well as chronic diseases such as atherosclerosis. Therefore, it is critically important to develop a regimen for generating memory monocytes in vitro in order to better define key monocyte memory states with diverse potentials for proliferation, differentiation, and activation, as well as underlying mechanisms. Here, we describe an efficient in vitro system to propagate a large number of highly purified murine memory monocytes through sustaining bone marrow-derived monocytes with macrophage colony-stimulating factor (M-CSF, 10 ng/mL)-containing medium, together with other polarization agents such as lipopolysaccharide (LPS) for a 5-day period. This method can yield high-purity monocytes, capable of exhibiting dynamic memory behaviors upon training with various polarizing agents., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Blood culture is the standard diagnostic method for typhoid and paratyphoid (enteric) fever in surveillance studies and clinical trials, but sensitivity is widely acknowledged to be suboptimal. We conducted a systematic review and meta-analysis to examine sources of heterogeneity across studies and quantified the effect of blood volume.Methods: We searched the literature to identify all studies that performed blood culture alongside bone marrow culture (a gold standard) to detect cases of enteric fever. We performed a meta-regression analysis to quantify the relationship between blood sample volume and diagnostic sensitivity. Furthermore, we evaluated the impact of patient age, antimicrobial use, and symptom duration on sensitivity.Results: We estimated blood culture diagnostic sensitivity was 0.59 (95% confidence interval [CI], 0.54-0.64) with significant between-study heterogeneity (I2, 76% [95% CI, 68%-82%]; P < .01). Sensitivity ranged from 0.51 (95% CI, 0.44-0.57) for a 2-mL blood specimen to 0.65 (95% CI, 0.58-0.70) for a 10-mL blood specimen, indicative of a relationship between specimen volume and sensitivity. Subgroup analysis showed significant heterogeneity by patient age and a weak trend towards higher sensitivity among more recent studies. Sensitivity was 34% lower (95% CI, 4%-54%) among patients with prior antimicrobial use and 31% lower after the first week of symptoms (95% CI, 19%-41%). There was no evidence of confounding by patient age, antimicrobial use, symptom duration, or study date on the relationship between specimen volume and sensitivity.Conclusions: The relationship between the blood sample volume and culture sensitivity should be accounted for in incidence and next-generation diagnostic studies. [ABSTRACT FROM AUTHOR]

Dendritic cells (DCs) are commonly generated from bone marrow (BM) progenitor cells with granulocyte-macrophage colony-stimulating factor (GM-CSF) alone or in combination with interleukin 4 (IL-4). These cells are often harvested post day 5, when they acquire maturation markers and can stimulate T cells. Apart from DCs, myeloid derived suppressor cells (MDSCs) are also found within these cultures. However, little is known about the functional characteristics of DCs and MDSCs before day 5. Herein, using a murine model, it is shown that early DCs and MDSCs, even in cultures with GM-CSF alone, upregulate fully maturation and activation surface molecules in response to the toll-like receptor 4 (TLR4) ligand lipopolysaccharide (LPS) stimulation. Despite initially displaying lower marker expression levels, these cells efficiently induced T cell stimulation and cytokine production. Interestingly, Gr-1int MDSCs increased their T cell co-stimulatory activity upon TLR4 stimulation. Additionally, early DCs and MDSCs exhibited differential endocytic capacity for viral sized nanoparticles and bacterial sized microparticles. DCs internalized both particle sizes, whilst MDSCs only internalized the larger microparticles, with reduced endocytic activity over time in the culture. These findings have unveiled an important role for the rapid initiation of productive immunity by GM-CSF, with promising implications for future vaccine and DC immunotherapy developments.

Mesangiogenic Progenitor Cells (MPCs) are human bone marrow-derived multipotent cells, isolated in vitro under selective culture conditions and shown to retain both mesengenic and angiogenic potential. MPCs also co-isolated with multipotent stromal cells (MSCs) when bone marrow primary cultures were set up for clinical applications, using human serum (HS) in place of fetal bovine serum (FBS). MPC culture purity (over 95%) is strictly dependent on HS supplementation with significant batch-to-batch variability. In the present paper we screened different sources of commercially available pooled human AB type serum (PhABS) for their ability to promote MPC production under selective culture conditions. As the majority of contaminating cells in MPC cultures were represented by MSC-like cells, we hypothesized a role by differentiating agents present in the sera. Therefore, we tested a number of growth factors (hGF) and found that higher concentrations of FGF-2, EGF, PDGF-AB, and VEGF-A as well as lower concentration of IGF-1 give sub-optimal MPC recovery. Gene expression analysis of hGF receptors was also carried out both in MSCs and MPCs, suggesting that FGF-2, EGF and PDGF-AB could act promoting MSC proliferation, while VEGF-A contribute to MSC-like cell contamination, triggering MPC differentiation. Here we demonstrated that managing hGF contents, together with applying specific receptors inhibitors (Erlotinib-HCl and Nintedanib), could significantly mitigate the batch-to-batch variability related to serum supplementation. These data represent a fundamental milestone in view of manufacturing MPC-based medicinal products.

Background: Various blood cells with different functions are produced in bone marrow in a process called hematopoiesis. Objective: The present study was conducted in order to evaluate the effect(s) of MS14, as an immunomodulator with natural origin (plant –marine) on the hematopoiesis. Methods: Eight 6-8 weeks old female BALB/c mice were divided to control group (receiving normal saline) and MS14 group (receiving MS14 at 100 mg/kg). Administration of MS14 and normal saline prolongs for five days, and then the mice were anaesthetized and killed. Smear of peripheral blood cells was provided and bone marrow cells were counted and cultured for 48 h. Erythropoietin (20 ng/ml) was added to half of samples. Results: 5 days administration of 100 mg/kg MS14 has augmented the percent of red blood cell of bone marrow (about 2 times). An increase (about 60%) in the percent of peripheral blood neutrophils has been observed as well. Erytroid colonies in bone marrow culture have been increased about 2 times in MS14 group e.i. the mean colony number in each well increased from 7 in control group to 14 in MS14 group and at the presence of erythropoietin from 13 colonies in control to 23 in MS14 group. Conclusion: According to increasing effect of MS14 on the number of erytroid colonies and percent of red blood cells, it can be concluded that hematopoietic processes not only does not adversely affected or inhibited by MS14 but could be significantly augmented when MS14 adminstered.

Introduction In acute myeloid leukemia ( AML), it has been shown that AML-derived cells often remain sensitive to autophagy-inducing stimuli, leading to the idea that harnessing the autophagy can be pertinent to AML cytotoxic therapy. Despite this promising notion, to date, there is no comprehensive study addressing autophagy-related genes expression status in AML. As a critical mediator, BECN1 influences the onset and advance of autophagy and several studies have pointed to the BECN1 recurrent allelic deletion and expression variation in a broad range of tumors. To explore this caveat, we chose this alteration-prone gene to investigate in our study. Methods We have analyzed the expression status of BECN1 in a series of 128 de novo AML patients using real-time quantitative polymerase chain reaction ( qRT- PCR). Results In our favorable subgroup, BECN1 expression did not alter ( P = 0.301), but in intermediate and unfavorable patients, we have had BECN1 low expression compared to the normal controls ( P = 0.008 and P < 0.001, respectively). We found evidence for the association of reduced expression of BECN1 with FLT3- ITD mutation (19 of 27 patients), monosomal karyotype (all of 11 patients), higher age, and WBC count. Conclusion Overall, remarkable association of reduced expression of BECN1 with FLT3- ITD mutation and monosomal karyotype and their functional relationship is interesting which should be addressed and verified in future studies. [ABSTRACT FROM AUTHOR]

In aplastic anaemia ( AA), haemopoietic activity is significantly reduced and generally attributed to failure of haemopoietic stem cells ( HSC) within the bone marrow ( BM). The regulation of haemopoiesis depends on the interaction between HSC and various cells of the BM microenvironment, including mesenchymal stromal cells ( MSC). MSC involvement in the functional restriction of HSC in AA is largely unknown and therefore, the physical and functional properties of AA MSC were studied in vitro. MSC were characterized by their phenotype and ability to form adherent stromal layers. The functional properties of AA MSC were assessed through proliferative, clonogenic and cross-over culture assays. Results indicate that although AA MSC presented typical morphology and distinctive mesenchymal markers, stromal formation was reduced, with 50% of BM samples failing to produce adherent layers. Furthermore, their proliferative and clonogenic capacity was markedly decreased ( P = 0·03 and P = 0·04 respectively) and the ability to sustain haemopoiesis was significantly reduced, as assessed by total cell proliferation ( P = 0·032 and P = 0·019 at Week 5 and 6, respectively) and clonogenic potential of HSC ( P = 0·02 at Week 6). It was concluded that the biological characteristics of AA MSC are different from those of control MSC and their in vitro haemopoiesis-supporting ability is significantly reduced. [ABSTRACT FROM AUTHOR]

Bone remodeling is a process consisting of bone formation and resorption. The present study compared the relative osteoclastic and osteoblastic potency of bone morphogenetic proteins (BMP)-2, -4, -5, -6, and -7 in primary murine bone marrow cultures. All five BMPs stimulated, to varying degree, formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells in a time- and protein concentration-dependent manner. The TRAP staining intensity correlated positively with the number of nuclei per TRAP-positive cell and the mRNA levels of colony-stimulating factor (CSF-1), receptor activator of nuclear factor κB ligand (RANKL), TRAP, and cathepsin K. Under osteogenic conditions, all five BMPs stimulated AP activity and mineralized bone nodule formation in a protein concentration-dependent manner in the same primary murine bone marrow cell culture system. These findings should be useful in designing treatment strategies for bone regeneration. [ABSTRACT FROM AUTHOR]

Summary: Objectives: Because of the suboptimal recovery rate of brucellae from blood, it has been proposed that cultures of bone marrow, liver tissue, and lymph nodes may improve the recovery rate of the organism. Data in support of these recommendations are limited and not clearly convincing, especially that of bone marrow culture. The main purpose of this work was to evaluate the roles of blood, bone marrow, liver, and lymph node cultures in the diagnosis of human brucellosis. Methods: Blood and bone marrow cultures were evaluated in parallel in 103 cases of human brucellosis using Castaneda''s biphasic technique. Simultaneous cultures of blood, bone marrow, liver, and lymph node aspirates were also carried out for 13 of these 103 cases. Results: Blood culture identified 47 (45.6%) cases and bone marrow culture identified 85 (82.5%) cases. Faster recovery of Brucella spp was accomplished with the bone marrow culture (2.8±0.7 days, p <0.05). When the results of cultures of blood and bone marrow were compared with each other in the 13 cases, it was found that bone marrow specimens could be sterile (six cases (46%)) when bacteremia was present, but Brucella melitensis was detected in liver aspirate in all these six bacteremic cases. Conclusions: Our data indicate that it is worthwhile practicing bone marrow culture by conventional biphasic technique for the definitive and rapid diagnosis of brucellosis; this is particularly the case in developing countries where diagnostic facilities by advanced technologies such as automated culture systems with PCR are not available. Bone marrow culturing would be a better gold standard in areas where antibiotic pretreatment is common. Also, adopting the practice of culturing liver/lymph node fluids may enhance bacterial isolation and aid in the establishment of a diagnosis of brucellosis in cases for whom blood and bone marrow cultures are negative. [Copyright &y& Elsevier]

Abstract: To better characterize acute myeloid leukemia (AML) development in non-obese diabetic (NOD)/severe combined immunodeficiency (SCID) mice, we transplanted samples from patients with AML or KG-1 and EOL-1 cell lines. We found 9/12 primary AML samples and both cell lines to engraft within 2–8 weeks, with 5–80% human cells in bone marrow. Compared with freshly isolated AML cells, percentages of human CD33+, CD38+, CD31+ CD13+ or CD15+ subpopulations increased after transplantation, whereas percentages of CD34+ cells decreased. Engrafted mice frequently showed expression of human endothelial cell markers. Thus, transplantation of human AML into NOD/SCID mice reveals expression of hematopoietic and endothelial differentiation markers. [Copyright &y& Elsevier]

Dendritic cells (DCs) are a heterogeneous population of cells of fundamental importance in initiating innate as well as specific immune responses. The identity and function of DCs in the cat are unknown, although they are likely pivotal in the response to infection. In this study, feline DCs were derived by 3–10-day culture of adherent blood mononuclear cells (PBMCs) and bone marrow mononuclear cells (BMMCs) in the presence of IL 4 and GM-CSF. BMMC consistently yielded a greater number of DCs than PBMC, and there were fewer macrophages than DC from both compartments. DCs expressed a distinct constellation of surface molecules, which included CD1a, CD1b, and CD1c, CD11b, CD14, and 2–3-fold higher levels of MHC class I and II molecules than co-cultured macrophages or fresh blood monocytes. DCs displayed typical cytoplasmic processes, limited non-specific esterase activity, and acquired antigen by phagocytosis, pinocytosis, and binding to specific receptors. Cytokine-exposed cells induced proliferation of allogeneic lymphocytes. Thus, the cells derived by these culture conditions had markers and functions analogous to immature myeloid DCs. Availability of feline DCs will enable investigation of their role in infectious disease and their potential therapeutic application. [Copyright &y& Elsevier]

Platelets are activated at fracture sites or upon the insertion of implants as a consequence of vascular disruption and secrete the contents of their granules into the developing hematoma. The regeneration of injured tissue requires bone remodeling and the resorbing activity of osteoclasts. To test our hypothesis that platelets can stimulate osteoclastogenesis, we examined the effects of supernatants released from thrombin-activated platelets on osteoclast-like cell formation in murine bone marrow cultures. Histochemical analysis indicated the presence of bone-resorbing, tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells. Transcripts that are characteristically expressed in native osteoclasts were increased in these cultures, as determined by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. The inhibition of both cyclooxygenases with indomethacin, as well as the addition of the cyclooxygenase-2 (COX-2)-selective antagonist, NS398, completely blocked osteoclast-like cell formation and decreased endogenous prostaglandin E2 production. Platelet-released supernatants stimulated the expression of receptor activator of NF-κB ligand (RANKL), whereas mRNA levels of osteoprotegerin (OPG) were decreased. The formation of osteoclast-like cells was prevented by recombinant OPG. Our results suggest that COX-2 activity is necessary for osteoclast-like cell formation in response to platelet-released supernatants, and that endogenously produced prostaglandin E2 can, in turn, increase the RANKL:OPG ratio, indicating that platelets can contribute to bone remodeling by stimulation of osteoclastogenesis. [Copyright &y& Elsevier]

Endothelin (ET)-1 induces proliferation of various cells including smooth muscle cells, fibroblasts, glomerular mesangial cells, endothelial cells and osteoblasts. ET-1 also stimulates synthesis of interleukin (IL)-6 in endothelial and bone marrow stromal cells of rat. It is well known that IL-6 modulates megakaryocytopoiesis. Some studies have indicated that megakaryocytes express both ET receptors and they are targets for ET. Therefore we planned to examine the effects of ET-1 on the growth of normal megakaryocytic cells in rat bone marrow primary cell culture. Bone marrow cells were cultured at 37°C, in an incubator atmosphere of 5% CO2 in air and 95% relative humidity for nine days. ET-1 at 10-7, 10-8 and 10-11 M, and control with saline were added at the beginning of the experiment protocol. At each day, plasma clots were stained using direct-coloring thiocholin method for acetylcholinesterase activity. Although 10-7 M ET-1 did not change the proliferation of megakaryocytic cells, this could be due to the presence of over crowded fibroblasts in the same environment. 10-8 M ET-1 stimulated megakaryocytic cell growth to 234% over the control on the fifth day. ET-1 at a concentration of 10-11 M also rised the megakaryocytic cell number significantly reaching up to 86% at the sixth day. Our results indicate that ET-1 may modulate the growth of megakaryocytic cells by an autocrine and/or paracrine action. [ABSTRACT FROM AUTHOR]

Enteric fever is one of the major clinical forms associated by Salmonella. Salmonella was detected unusually in the blood and bone marrow cultures of 19 year-old patient who was admitted to our hospital with high fever, abdominal pain, nausea, vomiting. However, antibody titer against Salmonella has not been detected in the serological tests. We present this case which was treated with antibioteraphy, namely ciprofloxacin and cefoperazone. [ABSTRACT FROM AUTHOR]

We describe the incidence and morphology of cellular aggregates which may develop in 8 day bone marrow (BM) cultures of patients with acute myeloid leukemia (AML). Aggregates formed in at least one BM culture from 50% (20/39) of the AML group. They developed irrespective of the patient's status (i.e. stages M1-M4), FAB type, and presence of colony stimulating factor (CSF). All aggregates were composed of macrophages, plasma cells, and cells of the myelocyte series surrounding a core of adipocytes and collagen fibrils. The percentage of blasts and promyelocytes in the plated BM aspirate governed the final composition of the aggregate. Patients in Stages M3 or M4 with FAB types M1 or M2 formed aggregates with a high proportion of myelocytic cells; aggregates of all other AML patients were composed predominantly of macrophages and plasmacytes. Aggregates appeared to form as a result of attraction of cells in the medium toward the stroma cell core. Furthermore, the development of aggregates in the absence of exogenous CSF, suggested that stromal cells excreted a factor with CSF-like activity. The results indicate that cellular aggregates in AML-BM cultures reflect the important role of BM stroma in creating microenvironments which enhance the development of hemopoietic stem cells. [ABSTRACT FROM AUTHOR]