Assessing the impact of COVID-19 on acute leukemia patients: a comparative analysis of hematological and biochemical parameters.

BACKGROUND: The impact of COVID-19 infection on the blood system remains to be investigated, especially with those encountering hematological malignancies. It was found that a high proportion of cancer patients are at an elevated risk of encountering COVID-19 infection. Leukemic patients are often suppressed and immunocompromised, which would impact the pathology following COVID-19 infection. Therefore, this research aims to bring valuable insight into the mechanism by which COVID-19 infection influences the hematological and biochemical parameters of patients with acute leukemia. METHODS: This retrospective investigation uses repeated measures to examine changes in hematological and biochemical parameters among patients with acute leukemia before and after COVID-19 infection at a major Saudi tertiary center. The investigation was conducted at the Ministry of National Guard-Health Affairs in Riyadh, Saudi Arabia, on 24 acute leukemia patients with COVID-19 between April 2020 and July 2023. The impact of COVID-19 on clinical parameters, comorbidities, and laboratory values was evaluated using data obtained from the electronic health records at four designated time intervals. The relative importance of comorbidities, testing preferences, and significant predictors of survival was ascertained. RESULTS: The majority of leukemic COVID-19-infected patients, primarily detected through PCR tests, were diagnosed with acute lymphoblastic leukemia (70.8%). The hematological and biochemical parameters exhibited stability, except for a brief increase in ALT and a sustained rise in AST. These changes were not statistically significant, and parameters remained normal at all time points. Additionally, an increase in monocyte count was shown at time point-3, as well as platelet counts at time point 2. CONCLUSION: While this study did not detect statistically significant effects of COVID-19 on biochemical and hematological parameters in acute leukemia patients, further investigation is needed to fully understand the potential adverse reactions and modifications following COVID-19 infection.

Paediatric COVID-19 Outcomes: Haematology Parameters, Mortality Rates, and Hospitalization Duration.

The global COVID-19 pandemic has strained healthcare systems around the globe, necessitating extensive research into the variables that affect patient outcomes. This study examines the relationships between key haematology parameters, duration of hospital stay (LOS), and mortality rates in COVID-19 cases in paediatric patients. Researchers analyse relationships between independent variables (COVID-19 status, age, sex) and dependent variables (mortality, LOS, coagulation parameters, WBC count, RBC parameters) using multivariate regression models. Although the R-square values (0.6-3.7%) indicate limited explanatory power, coefficients with statistical significance establish the impact of independent variables on outcomes. Age emerges as a crucial predictor of mortality; the mortality rate decreases by 1.768% per age group. Both COVID-19 status and age have an inverse relationship with length of stay, emphasising the milder hospitalisation of children. Platelet counts decline with age and male gender, potentially revealing the influence of COVID-19 on haematological markers. There are significant correlations between COVID-19 status, age, gender and coagulation measures. Lower prothrombin time and D-dimer concentrations in elder COVID-19 patients are indicative of distinct coagulation profiles. WBC and RBC parameters exhibit correlations with variables: COVID-19-positive patients have lower WBC counts, whereas male COVID-19-positive patients have higher RBC counts. In addition, correlations exist between independent variables and the red cell distribution width, mean corpuscular volume, and mean corpuscular haemoglobin. However, there is no correlation between mean corpuscular haemoglobin concentration and outcomes, indicating complex interactions between haematological markers and outcomes. In essence, this study underlines the importance of age in COVID-19 mortality, provides novel insights into platelet counts, and emphasises the complexity of the relationships between haematological parameters and disease outcomes.

Effect of Achillea fragrantissima Extract on Excision Wound Biofilms of MRSA and Pseudomonas aeruginosa in Diabetic Mice.

Achillea fragrantissima, a desert plant commonly known as yarrow, is traditionally used as an antimicrobial agent in folklore medicine in Saudi Arabia. The current study was undertaken to determine its antibiofilm activity against methicillin-resistant Staphylococcus aureus (MRSA) and multi-drug-resistant Pseudomonas aeruginosa (MDR-P. aeruginosa) using in vitro and in vivo studies. A biofilm model induced through an excision wound in diabetic mice was used to evaluate its effect in vivo. The skin irritation and cytotoxic effects of the extract were determined using mice and HaCaT cell lines, respectively. The Achillea fragrantissima methanolic extract was analyzed with LC-MS to detect different phytoconstituents, which revealed the presence of 47 different phytoconstituents. The extract inhibited the growth of both tested pathogens in vitro. It also increased the healing of biofilm-formed excision wounds, demonstrating its antibiofilm, antimicrobial, and wound-healing action in vivo. The effect of the extract was concentration-dependent, and its activity was stronger against MRSA than MDR-P. aeruginosa. The extract formulation was devoid of a skin irritation effect in vivo and cytotoxic effect on HaCaT cell lines in vitro.

A Simple In-Vivo Method for Evaluation of Antibiofilm and Wound Healing Activity Using Excision Wound Model in Diabetic Swiss Albino Mice.

The study developed a simple and inexpensive method to induce biofilm formation in-vivo for the evaluation of the antibiofilm activity of pharmacological agents using Swiss albino mice. Animals were made diabetic using streptozocin and nicotinamide. A cover slip containing preformed biofilm along with MRSA culture was introduced into the excision wound in these animals. The method was effective in developing biofilm on the coverslip after 24 h incubation in MRSA broth which was confirmed by microscopic examination and a crystal violet assay. Application of preformed biofilm along with microbial culture induced a profound infection with biofilm formation on excision wounds in 72 h. This was confirmed by macroscopic, histological, and bacterial load determination. Mupirocin, a known antibacterial agent effective against MRSA was used to demonstrate antibiofilm activity. Mupirocin was able to completely heal the excised wounds in 19 to 21 days while in the base-treated group, healing took place between 30 and 35 days. The method described is robust and can be reproduced easily without the use of transgenic animals and sophisticated methods such as confocal microscopy.

C/EBPß promotes the expression of atrophy-inducing factors by tumours and is a central regulator of cancer cachexia.

BACKGROUND: CCAAT/enhancer-binding protein ß (C/EBPß) is a transcription factor whose high expression in human cancers is associated with tumour aggressiveness and poor outcomes. Most advanced cancer patients will develop cachexia, characterized by loss of skeletal muscle mass. In response to secreted factors from cachexia-inducing tumours, C/EBPß is stimulated in muscle, leading to both myofibre atrophy and the inhibition of muscle regeneration. Involved in the regulation of immune responses, C/EBPß induces the expression of many secreted factors, including cytokines. Because tumour-secreted factors drive cachexia and aggressive tumours have higher expression of C/EBPß, we examined a potential role for C/EBPß in the expression of tumour-derived cachexia-inducing factors. METHODS: We used gain-of-function and loss-of-function approaches in vitro and in vivo to evaluate the role of tumour C/EBPß expression on the secretion of cachexia-inducing factors. RESULTS: We report that C/EBPß overexpression up-regulates the expression of 260 secreted protein genes, resulting in a secretome that inhibits myogenic differentiation (-31%, P < 0.05) and myotube maturation [-38% (fusion index) and -25% (myotube diameter), P < 0.05]. We find that knockdown of C/EBPß in cachexia-inducing Lewis lung carcinoma cells restores myogenic differentiation (+25%, P < 0.0001) and myotube diameter (+90%, P < 0.0001) in conditioned medium experiments and, in vivo, prevents muscle wasting (-51% for small myofibres vs. controls, P < 0.01; +140% for large myofibres, P < 0.01). Conversely, overexpression of C/EBPß in non-cachectic tumours converts their secretome into a cachexia-inducing one, resulting in reduced myotube diameter (-41%, P < 0.0001, EL4 model) and inhibition of differentiation in culture (-26%, P < 0.01, EL4 model) and muscle wasting in vivo (+98% small fibres, P < 0.001; -76% large fibres, P < 0.001). Comparison of the differently expressed transcripts coding for secreted proteins in C/EBPß-overexpressing myoblasts with the secretome from 27 different types of human cancers revealed ~18% similarity between C/EBPß-regulated secreted proteins and those secreted by highly cachectic tumours (brain, pancreatic, and stomach cancers). At the protein level, we identified 16 novel secreted factors that are present in human cancer secretomes and are up-regulated by C/EBPß. Of these, we tested the effect of three factors (SERPINF1, TNFRSF11B, and CD93) on myotubes and found that all had atrophic potential (-33 to -36% for myotube diameter, P < 0.01). CONCLUSIONS: We find that C/EBPß is necessary and sufficient to induce the secretion of cachexia-inducing factors by cancer cells and loss of C/EBPß in tumours attenuates muscle atrophy in an animal model of cancer cachexia. Our findings establish C/EBPß as a central regulator of cancer cachexia and an important therapeutic target.

From quiescence to repair: C/EBPß as a regulator of muscle stem cell function in health and disease.

CCAAT/Enhancer Binding protein beta (C/EBPß) is a transcriptional regulator involved in numerous physiological processes. Herein, we describe a role for C/EBPß as a regulator of skeletal muscle stem cell function. In particular, C/EBPß is expressed in muscle stem cells in healthy muscle where it inhibits myogenic differentiation. Downregulation of C/EBPß expression at the protein and transcriptional level allows for differentiation. Persistence of C/EBPß promotes stem cell self-renewal and C/EBPß expression is required for mitotic quiescence in this cell population. As a critical regulator of skeletal muscle homeostasis, C/EBPß expression is stimulated in pathological conditions such as cancer cachexia, which perturbs muscle regeneration and promotes myofiber atrophy in the context of systemic inflammation. C/EBPß is also an important regulator of cytokine expression and immune response genes, a mechanism by which it can influence muscle stem cell function. In this viewpoint, we describe a role for C/EBPß in muscle stem cells and propose a functional intersection between C/EBPß and NF-kB action in the regulation of cancer cachexia.

SMAD2 promotes myogenin expression and terminal myogenic differentiation.

SMAD2 is a transcription factor, the activity of which is regulated by members of the transforming growth factor ß (TGFß) superfamily. Although activation of SMAD2 and SMAD3 downstream of TGFß or myostatin signaling is known to inhibit myogenesis, we found that SMAD2 in the absence of TGFß signaling promotes terminal myogenic differentiation. We found that, during myogenic differentiation, SMAD2 expression is induced. Knockout of SMAD2 expression in primary myoblasts did not affect the efficiency of myogenic differentiation but produced smaller myotubes with reduced expression of the terminal differentiation marker myogenin. Conversely, overexpression of SMAD2 stimulated myogenin expression, and enhanced both differentiation and fusion, and these effects were independent of classical activation by the TGFß receptor complex. Loss of Smad2 in muscle satellite cells in vivo resulted in decreased muscle fiber caliber and impaired regeneration after acute injury. Taken together, we demonstrate that SMAD2 is an important positive regulator of myogenic differentiation, in part through the regulation of Myog.

CCAAT/enhancer-binding protein beta promotes muscle stem cell quiescence through regulation of quiescence-associated genes.

Regeneration of skeletal muscle depends on resident muscle stem cells called satellite cells that in healthy, uninjured muscle remain quiescent (noncycling). After activation and expansion of satellite cells postinjury, satellite cell numbers return to uninjured levels and return to mitotic quiescence. Here, we show that the transcription factor CCAAT/enhancer-binding protein beta (C/EBPß) is required to maintain quiescence of satellite cells in uninjured muscle. We show that C/EBPß is expressed in quiescent satellite cells in vivo and upregulated in noncycling myoblasts in vitro. Loss of C/EBPß in satellite cells promotes their premature exit from quiescence resulting in spontaneous activation and differentiation of the stem cell pool. Forced expression of C/EBPß in myoblasts inhibits proliferation by upregulation of 28 quiescence-associated genes. Furthermore, we find that caveolin-1 is a direct transcriptional target of C/EBPß and is required for cell cycle exit in muscle satellite cells expressing C/EBPß. The induction of mitotic quiescence is considered necessary for the long-term maintenance of adult stem cell populations with dysregulation driving increased differentiation of progenitors and depletion of the stem cell pool. Our findings place C/EBPß as an important transcriptional regulator of muscle satellite cell quiescence.

Contaminating reactivity of a monoclonal CCAAT/Enhancer Binding Protein ß antibody in differentiating myoblasts.

OBJECTIVE: CCAAT/Enhancer Binding proteins (C/EBPs) are transcription factors involved in the regulation of a variety of cellular processes. We used the Abcam Recombinant Anti-C/EBP beta antibody (E299) to detect C/EBPß expression during myogenesis. Though the antibody is monoclonal, and the immunogen used is highly specific to C/EBPß, we identified an intense band at 23 kDa on western blot that did not correspond to any of the known isoforms of C/EBPß, or family members predicted to cross-react. Absent in myoblast cells overexpressing C/EBPß, the band was present when C/EBPß was knocked down, confirming specificity for a protein other than C/EBPß. The objective of this work was to identify the contaminating reactivity. RESULTS: We performed immunoprecipitation followed by mass spectrometry to identified myosin light chain 4 (MYL4) as the unknown band, suggesting that the Abcam monoclonal antibody directed against C/EBPß is not pure, but contains a contaminating antibody against MYL4. Caution should be used when working in cells lines that express MYL4 to not confound the detection of MYL4 with that of C/EBPß isoforms.

CCAAT/Enhancer Binding Protein ß inhibits myogenic differentiation via ID3.

Myogenesis is regulated by the coordinated expression of muscle regulatory factors, a family of transcription factors that includes MYOD, MYF5, myogenin and MRF4. Muscle regulatory factors are basic helix-loop-helix transcription factors that heterodimerize with E proteins to bind the regulatory regions of target genes. Their activity can be inhibited by members of the Inhibitor of DNA binding and differentiation (ID) family, which bind E-proteins with high affinity, thereby preventing muscle regulatory factor-dependent transcriptional responses. CCAAT/Enhancer Binding protein beta (C/EBPß) is a transcription factor expressed in myogenic precursor cells that acts to inhibit myogenic differentiation, though the mechanism remains poorly understood. We identify Id3 as a novel C/EBPß target gene that inhibits myogenic differentiation. Overexpression of C/EBPß stimulates Id3 mRNA and protein expression, and is required for C/EBPß-mediated inhibition of myogenic differentiation. Misexpression of C/EBPß in myogenic precursors, such as in models of cancer cachexia, prevents the differentiation of myogenic precursors and we show that loss of Id3 rescues differentiation under these conditions, suggesting that the stimulation of Id3 expression by C/EBPß is an important mechanism by which C/EBPß inhibits myogenic differentiation.

CCAAT/enhancer binding protein ß is required for satellite cell self-renewal.

BACKGROUND: Postnatal growth and repair of skeletal muscle relies upon a population of quiescent muscle precursor cells, called satellite cells that can be activated to proliferate and differentiate into new myofibers, as well as self-renew to replenish the satellite cell population. The balance between differentiation and self-renewal is critical to maintain muscle tissue homeostasis, and alterations in this equilibrium can lead to chronic muscle degeneration. The transcription factor CCAAT/enhancer binding protein beta (C/EBPß) is expressed in Pax7+ satellite cells of healthy muscle and is downregulated during myoblast differentiation. Persistent expression of C/EBPß upregulates Pax7, inhibits MyoD, and blocks myogenic differentiation. METHODS: Using genetic tools to conditionally abrogate C/EBPß expression in Pax7+ cells, we examined the role of C/EBPß in self-renewal of satellite cells during muscle regeneration. RESULTS: We find that loss of C/EBPß leads to precocious differentiation at the expense of self-renewal in primary myoblast and myofiber cultures. After a single muscle injury, C/EBPß-deficient satellite cells fail to self-renew resulting in a reduction of satellite cells available for future rounds of regeneration. After a second round of injury, muscle regeneration is impaired in C/EBPß conditional knockout mice compared to wild-type control mice. We find that C/EBPß can regulate Notch2 expression and that restoration of Notch activity in myoblasts lacking C/EBPß prevents precocious differentiation. CONCLUSIONS: These findings demonstrate that C/EBPß is a novel regulator of satellite cell self-renewal during muscle regeneration acting at least in part through Notch2.

Retinoid X Receptor-selective Signaling in the Regulation of Akt/Protein Kinase B Isoform-specific Expression.

The differentiation and fusion of myoblasts into mature myotubes are complex processes responding to multiple signaling pathways. The function of Akt/PKB is critical for myogenesis, but less is clear as to the regulation of its isoform-specific expression. Bexarotene is a drug already used clinically to treat cancer, and it has the ability to enhance the commitment of embryonic stem cells into skeletal muscle lineage. Whereas bexarotene regulates fundamental biological processes through retinoid X receptor (RXR)-mediated gene expression, molecular pathways underlying its positive effects on myogenesis remain unclear. In this study, we have examined the signaling pathways that transmit bexarotene action in the context of myoblast differentiation. We show that bexarotene promotes myoblast differentiation and fusion through the activation of RXR and the regulation of Akt/PKB isoform-specific expression. Interestingly, bexarotene signaling appears to correlate with residue-specific histone acetylation and is able to counteract the detrimental effects of cachectic factors on myogenic differentiation. We also signify an isoform-specific role for Akt/PKB in RXR-selective signaling to promote and to retain myoblast differentiation. Taken together, our findings establish the viability of applying bexarotene in the prevention and treatment of muscle-wasting disorders, particularly given the lack of drugs that promote myogenic differentiation available for potential clinical applications. Furthermore, the model of bexarotene-enhanced myogenic differentiation will provide an important avenue to identify additional genetic targets and specific molecular interactions that we can study and apply for the development of potential therapeutics in muscle regeneration and repair.