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Myeloid-Derived Suppressor Cells (MDSC) in Melanoma Patients Treated with Anti-PD-1 Immunotherapy
Myeloid-derived suppressor cells (MDSC) are a subset of immature myeloid cells with suppressive activity well described in the context of cancer. They inhibit anti-tumour immunity, promote metastasis formation and can lead to immune therapy resistance. In a retrospective study, blood probes of 46 advanced melanoma patients were analysed before the first administration of anti-PD-1 immunotherapy and in the third month of treatment for MDSC, immature monocytic (ImMC), monocytic MDSC (MoMDSC) and granulocytic MDSC (GrMDSC) by multi-channel flow cytometry. Cell frequencies were correlated with response to immunotherapy, progression-free survival (PFS) and lactate dehydrogenase (LDH) serum level. Responders to anti-PD-1 therapy had higher MoMDSC levels (4.1 ± 1.2%) compared to non-responders (3.0 ± 1.2%) (p = 0.0333) before the first administration of anti-PD-1. No significant changes in MDSCs frequencies were observed in the groups of patients before and in the third month of therapy. The cut-off values of MDSCs, MoMDSCs, GrMDSCs and ImMCs for favourable 2- and 3-year PFS were established. Elevated LDH level is a negative prognostic factor of response to the treatment and is related to an elevated ratio of GrMDSCs and ImMCs level compared to patients’ LDH level below the cut-off. Our data may provide a new perspective for more careful consideration of MDSCs, and specially MoMDSCs, as a tool for monitoring the immune status of melanoma patients. Changes in MDSC levels may have a potential prognostic value, however a correlation with other parameters must be established.
A Clinical Outcome of the Anti-PD-1 Therapy of Melanoma in Polish Patients Is Mediated by Population-Specific Gut Microbiome Composition
The gut microbiota is considered a key player modulating the efficacy of immune checkpoint inhibitor therapy. The study investigated the association between the response to anti-PD-1 therapy and the baseline gut microbiome in a Polish cohort of melanoma patients, alongside selected agents modifying the microbiome. Sixty-four melanoma patients enrolled for the anti-PD-1 therapy, and ten healthy subjects were recruited. The response to the treatment was assessed according to the response evaluation criteria in solid tumors, and patients were classified as responders or non-responders. The association between selected extrinsic factors and response was investigated using questionnaire-based analysis and the metataxonomics of the microbiota. In the responders, the Bacteroidota to Firmicutes ratio was higher, and the richness was decreased. The abundance of Prevotella copri and Bacteroides uniformis was related to the response, whereas the non-responders’ gut microbiota was enriched with Faecalibacterium prausnitzii and Desulfovibrio intestinalis and some unclassified Firmicutes. Dietary patterns, including plant, dairy, and fat consumption as well as gastrointestinal tract functioning were significantly associated with the therapeutic effects of the therapy. The specific gut microbiota along with diet were found to be associated with the response to the therapy in the population of melanoma patients.
Nonhemolytic Listeria monocytogenes - Prevalence Rate, Reasons Underlying Atypical Phenotype, and Methods for Accurate Hemolysis Assessment
Listeria monocytogenes is a foodborne pathogen that typically presents β-hemolytic activity. However, there are literature reports indicating that L. monocytogenes strains are sometimes nonhemolytic or their zones of hemolysis are perceivable only after removal of the colonies from the agar plate. Nonhemolytic L. monocytogenes are most commonly encountered in food products, but some have also been detected in clinical samples. Usually, atypical bacteria of this species belong to serotype 1/2a. Mutations of the prfA gene sequence are the most common reason for changed phenotype, and mutations of the hly gene are the second most common cause. There are also reports that the methodology used for detecting hemolysis may influence the results. Sheep or horse blood, although most commonly used in modern studies, may not allow for the production of clear hemolytic zones on blood agar, whereas other types of blood (guinea pig, rabbit, piglet, and human) are more suitable according to some studies. Furthermore, the standard blood agar plate technique is less sensitive than its modifications such as bilayer or top-layer (overlay) techniques. The microplate technique (employing erythrocyte suspensions) is probably the most informative when assessing listerial hemolysis and is the least susceptible to subjective interpretation.
Characteristics of Intestinal Barrier State and Immunoglobulin-Bound Fraction of Stool Microbiota in Advanced Melanoma Patients Undergoing Anti-PD-1 Therapy
The gut microbiota is recognized as one of the extrinsic factors that modulate the clinical outcomes of immune checkpoint inhibitors (ICIs), such as inhibitors targeting programmed cell death protein 1 (PD-1), in cancer patients. However, the link between intestinal barrier, which mutually interacts with the gut microbiota, and therapeutic effects has not been extensively studied so far. Therefore, the primary goal of this study was to investigate the relationship between intestinal barrier functionality and clinical outcomes of anti-PD-1 therapy in patients with advanced melanoma. Fecal samples were collected from 64 patients before and during anti-PD-1 therapy. The levels of zonulin, calprotectin, and secretory immunoglobulin A (SIgA), which reflect intestinal permeability, inflammation, and immunity, respectively, were measured in fecal samples (n = 115) using an Enzyme-Linked Immunosorbent Assay (ELISA). Moreover, the composition of the immunoglobulin (Ig)-bound (n = 108) and total stool microbiota (n = 117) was determined by the V3–V4 region of 16S rRNA gene sequencing. ELISA indicated a higher baseline concentration of fecal SIgA in patients with favorable clinical outcomes than those with unfavorable ones. Moreover, high baseline concentrations of intestinal barrier state biomarkers correlated with survival outcomes. In the cases of fecal zonulin and fecal SIgA, there was a positive correlation, while in the case of fecal calprotectin, there was a negative correlation. Furthermore, there were differences in the microbial profiles of the Ig-bound stool microbiota between patients with favorable and unfavorable clinical outcomes and their changes during treatment. Collectively, these findings indicate an association between intestinal barrier functionality and clinical outcomes of anti-PD-1 therapy in advanced melanoma patients.
Characteristics of Intestinal Barrier State and Immunoglobulin-Bound Fraction of Stool Microbiota in Advanced Melanoma Patients Undergoing Anti-PD-1 Therapy
The gut microbiota is recognized as one of the extrinsic factors that modulate the clinical outcomes of immune checkpoint inhibitors (ICIs) in cancer patients. However, the role of the intestinal barrier, which mutually interacts with the gut microbiota, in shaping anti-cancer immune responses has not been extensively studied so far. Therefore, the primary goal of our study was to investigate the relationship between intestinal barrier functionality and clinical outcomes of anti-PD-1 therapy in patients with advanced melanoma. Fecal samples were collected from 64 patients before and during anti-PD-1 therapy. The levels of zonulin, calprotectin, and secretory immunoglobulin A (SIgA), which reflect intestinal permeability, inflammation, and immunity, respectively, were measured in fecal samples (n = 115) using ELISA. Moreover, the composition of the immunoglobulin (Ig)-bound (n = 108) and total stool microbiota (n = 117) was determined by the V3-V4 region of 16S rRNA gene sequencing. ELISA indicated a higher baseline concentration of fecal SIgA in patients with favorable clinical outcomes than those with unfavorable ones. Moreover, high baseline concentrations of intestinal barrier state biomarkers correlated with survival outcomes. In the cases of fecal zonulin and fecal SIgA, there was a positive correlation, while in the case of fecal calprotectin, a negative correlation. Furthermore, there were differences in the microbial profiles of the Ig-bound stool microbiota between patients with favorable and unfavorable clinical outcomes and their changes during treatment. Collectively, intestinal barrier functionality was associated with clinical outcomes of anti-PD-1 therapy in advanced melanoma patients. Future studies are warranted to elucidate whether intestinal barrier modification could improve ICI efficacy and estimate the clinical value and utility of biomarkers reflecting its state.
Impact of isoflavones and Lactobacillus acidophilus on the fecal microbiology status in healthy female rats
Effects of isoflavone and probiotic intake on calcium transport and bone metabolism biomarkers in female rats
AbstractCalcium is essential for maintaining bone health as it contributes to bone formation, remodeling, strength, and density. This study investigated the effect of isoflavones and probiotics on calcium transporters' gene expression, serum calcium levels, and bone metabolism biomarkers in healthy female rats. Forty‐eight female Wistar rats were classified into six groups. Bone metabolism biomarkers (pyridinoline, deoxypyridinoline, parathyroid hormone, and osteocalcin) and serum calcium levels were measured by enzyme‐linked immunosorbent assay (ELISA) and atomic absorption spectroscopy (AAS), respectively. Gene expression of calcium transporters (Trpv5 and Trpv6) was evaluated in duodenum and jejunum tissue samples using quantitative polymerase chain reaction (qPCR). Trpv5 and Trpv6, epithelial calcium channels, play a crucial role in calcium transport and homeostasis in the body. The study consisted of a1‐week adaptation period for the rats to adjust to the controlled conditions, followed by an 8‐week intervention phase. The daidzein and genistein group showed a significant increase in the gene expression of the Trpv6 transporter in the duodenum and a marked decrease in serum pyridinoline levels compared to the control group. The tempeh and soybean groups showed a significant decrease in the gene expression of the Trpv5 calcium transporter in the jejunum. However, no significant influence of the Lactobacillus acidophilus diet on calcium transport and bone metabolism biomarkers was observed in the L. acidophilus group. The correlation analysis showed a significant positive relationship between serum calcium, bone metabolism biomarkers, and calcium transporters. In conclusion, our study demonstrates that the daidzein and genistein diet improves calcium transport in the duodenum and reduces pyridinoline serum concentrations, while tempeh and soybean diets reduce calcium transport in the jejunum. However, the combination of daidzein, genistein, and L. acidophilus did not demonstrate a synergistic effect on calcium transport and bone metabolism, suggesting that further investigations are needed to elucidate their potential interactions.
β-glucuronidase activity is associated with carbohydrate metabolism but not with androgen status in overweight and obese women with polycystic ovary syndrome
Phage-Based Control of Listeria innocua in the Food Industry: A Strategy for Preventing Listeria monocytogenes Persistence in Biofilms
Listeria innocua, though considered non-pathogenic, frequently coexists with Listeria monocytogenes in industrial environments, aiding its survival in biofilms. These biofilms pose a significant challenge in food processing facilities, as they protect bacteria from disinfectants and facilitate their spread. The aim of this review was to identify bacteriophages as a promising method for eliminating Listeria biofilms from the food industry. Lytic bacteriophages show great potential in combating Listeria biofilms. Commercially available products, such as PhageGuard Listex™ (P100) (Micreos Food Safety, Wageningen, The Netherlands), effectively reduce both L. monocytogenes and L. innocua in food products and on production surfaces. Additionally, phage-derived enzymes, such as endolysins, can degrade biofilms, eliminating bacteria without compromising food quality. The following article highlights that although bacteriophages present a promising biocontrol method, further research is necessary to assess their long-term effectiveness, particularly regarding bacterial resistance. To maximize efficacy, a combination of strategies such as phage cocktails and disinfectants is recommended to enhance biofilm eradication and minimize food contamination risks.
Experimental Capacity of Human Fecal Microbiota to Degrade Fiber and Produce Short-Chain Fatty Acids Is Associated with Diet Quality and Anthropometric Parameters
Exploring correlations between gut mycobiome and lymphocytes in melanoma patients undergoing anti-PD-1 therapy
Abstract Research has shown that the microbiome can influence how the immune system responds to melanoma cells, affecting the course of the disease and the outcome of the therapy. Here, we used the metagenomic approach and flow cytometry analyses of blood cells to discover correlations between gut fungi of metastatic melanoma patients enrolled in anti-PD-1 therapy and lymphocytes in their blood. We analyzed the patterns of associations before the first administration of anti-PD-1 therapy (BT, n = 61) and in the third month of the therapy (T3, n = 37), allowing us to track changes during treatment. To understand the possible impact of gut fungi on the efficacy of anti-PD-1 therapy, we analyzed the associations in clinical beneficiaries (CB, n = 37) and non-beneficiaries (NB, n = 24), as well as responders (R, n = 28) and non-responders (NR, n = 33). Patients with LDH < 338 units/L, overall survival (OS) > 12, CB, as well as R, had lower levels of Shannon diversity (p = 0.02, p = 0.05, p = 0.05, and p = 0.03, respectively). We found that the correlation pattern between intestinal fungi and lymphocytes was specific to the type of response, positive or negative. When comparing CB and NB groups, correlations with opposite directions were detected for C. albicans, suggesting a response-specific immune reaction. For CB, M. restricta exhibited a set of correlations with different types of lymphocytes, with prevalent positive correlations, suggesting a robust immune response in the CB group. This result extends our former research, where M. restricta and C. albicans were associated with an increased risk of melanoma progression and a poorer response to anti-PD-1 treatment.
Antioxidant Activity, Probiotic Survivability, and Sensory Properties of a Phenolic-Rich Pulse Snack Bar Enriched with Lactiplantibacillus plantarum
Charakterystyka frakcji mikrobioty jelitowej związanej z wydzielniczymi immunoglobulinami A (SlgA) u pacjentów z zaawansowanym czerniakiem poddanych immunoterapii anty-PD-1
Lactiplantibacillus plantarum, Duodenal Hydroxyphenyllactic Acid and Iron: Insights from a Rat Model of a High-Fat Iron-Deficient Diet
The standardisation of the approach to metagenomic human gut analysis: from sample collection to microbiome profiling
AbstractIn recent years, the number of metagenomic studies increased significantly. Wide range of factors, including the tremendous community complexity and variability, is contributing to the challenge in reliable microbiome community profiling. Many approaches have been proposed to overcome these problems making hardly possible to compare results of different studies. The significant differences between procedures used in metagenomic research are reflected in a variation of the obtained results. This calls for the need for standardisation of the procedure, to reduce the confounding factors originating from DNA isolation, sequencing and bioinformatics analyses in order to ensure that the differences in microbiome composition are of a true biological origin. Although the best practices for metagenomics studies have been the topic of several publications and the main aim of the International Human Microbiome Standard (IHMS) project, standardisation of the procedure for generating and analysing metagenomic data is still far from being achieved. To highlight the difficulties in the standardisation of metagenomics methods, we thoroughly examined each step of the analysis of the human gut microbiome. We tested the DNA isolation procedure, preparation of NGS libraries for next-generation sequencing, and bioinformatics analysis, aimed at identifying microbial taxa. We showed that the homogenisation time is the leading factor impacting sample diversity, with the recommendation for a shorter homogenisation time (10 min). Ten minutes of homogenisation allows for better reflection of the bacteria gram-positive/gram-negative ratio, and the obtained results are the least heterogenous in terms of beta-diversity of samples microbial composition. Besides increasing the homogenisation time, we observed further potential impact of the library preparation kit on the gut microbiome profiling. Moreover, our analysis revealed that the choice of the library preparation kit influences the reproducibility of the results, which is an important factor that has to be taken into account in every experiment. In this study, a tagmentation-based kit allowed for obtaining the most reproducible results. We also considered the choice of the computational tool for determining the composition of intestinal microbiota, with Kraken2/Bracken pipeline outperforming MetaPhlAn2 in our in silico experiments. The design of an experiment and a detailed establishment of an experimental protocol may have a serious impact on determining the taxonomic profile of the intestinal microbiome community. Results of our experiment can be helpful for a wide range of studies that aim to better understand the role of the gut microbiome, as well as for clinical purposes.
A Clinical Outcome of the anti-PD-1 Therapy of Melanoma in Polish Patients Is Mediated by Population-Specific Gut Microbiome Composition
Gut microbiota is considered a key player modulating the efficacy of immune checkpoint inhibitor therapy. The study investigated the association between response to the anti-PD-1 therapy and the baseline gut microbiome in the Polish cohort of melanoma patients, alongside selected agents modifying the microbiome. Sixty-four melanoma patients enrolled for the anti-PD-1 therapy and 10 healthy subjects were recruited. Response to the treatment was assessed according to the response evaluation criteria in solid tumors, and patients were classified as responders or non-responders. The association between selected extrinsic factors and response was investigated using questionnaire-based analysis, and metataxonomics of the microbiota. The Bacteroidota to Firmicutes ratio was higher, and the richness was decreased in the responders. The abundance of Prevotella copri and Bacteroides uniformis was related to the response, whereas non-responder gut microbiota was enriched with Faecalibacterium prausnitzii and Desulfovibrio intestinalis, and some unclassified Firmicutes. Dietary patterns, including plant, dairy, and fat consumption, but also gastrointestinal tract functioning were significantly associated with the therapeutic effects of the therapy. The specific gut microbiota alongside diet were found associated with response to the therapy in the Polish population of melanoma patients.
Influence of isoflavones and probiotics on magnesium status in healthy female rats
Isoflavones and probiotics are promising nutrients for bone health, and magnesium (Mg) is essential for bone metabolism. This study aimed to determine the effects of daidzein, genistein and Lactobacillus acidophilus on the Mg status of healthy female rats. Forty-eight rats were randomly assigned to six groups, with the control group receiving a standard diet (AIN 93M). The remaining groups were fed the same diet with added ingredients such as tempeh flour; soy flour; pure daidzein and genistein; L. acidophilus or a combination of daidzein, genistein, and L. acidophilus. Tissue samples were collected after the eight-week intervention, and Mg concentrations were analysed using flame atomic absorption spectrometry. Myeloid and erythroid cells were determined using the haematoxylin and eosin bone staining method. Statistical analyses were conducted using one-way ANOVA with Tukey’s post hoc test and Pearson’s correlation coefficient. The threshold for significance was p < 0.05. Compared with the control group, adding tempeh to the diet of rats resulted in significant changes in Mg concentrations in various tissues, with a decrease in the kidneys and an increase in the fur. Although not statistically significant compared to the control group, the tempeh group showed increased Mg concentrations in the femur and spleen. The myeloid-to-erythroid cell ratio did not differ significantly among groups, but all intervention groups showed higher ratios than the control group. A strong negative correlation was observed between Mg concentrations in the kidneys and fur. Conversely, a positive correlation was identified between Mg concentrations in the pancreas and fur. Daily consumption of tempeh may improve Mg status in the organism. Intake of pure daidzein, genistein, or probiotic seems to have no effect on Mg concentrations in healthy rats.
Listeria monocytogenes Isolates from Meat Products and Processing Environment in Poland Are Sensitive to Commonly Used Antibiotics, with Rare Cases of Reduced Sensitivity to Ciprofloxacin
Antibiotic resistance is a global health problem, causing not only an increased mortality rate of bacterial infections but also economic losses due to, among other reasons, the need for longer hospital stays. Listeria monocytogenes is one of the foodborne pathogens with the ability to induce a serious illness called listeriosis, with approximately 20–30% fatal outcomes. The treatment regimen of listeriosis in humans includes the administration of antibiotics (in most cases, ampicillin or trimethoprim with sulfamethoxazole in case of allergies to β-lactams), so the resistance of this pathogen to antibiotics can potentially lead to increased mortality. The antibiotic sensitivity status of n = 153 L. monocytogenes isolates originating from meat food samples (raw and processed) and meat-processing environment (both contacting and non-contacting with food) collected between October 2020 and November 2021 in Poland was examined in this study. Susceptibility to antibiotics was determined using the disc diffusion method on Mueller–Hinton agar plates. All collected samples were susceptible to 9 antibiotics: ampicillin (10 µg), chloramphenicol (30 µg), erythromycin (15 µg), gentamicin (10 µg), penicillin (10 IU), streptomycin (10 µg), sulfamethoxazole/trimethoprim (1.25/23.75 µg), tetracycline (30 µg) and vancomycin (30 µg). Some of the isolates (n = 10; 6.5%) showed reduced susceptibility to ciprofloxacin (5 µg), which was classified as an intermediate response. All these ten isolates were collected from surfaces contacting with food in food-processing facilities.
Host Factors Associated with Gut Mycobiome Structure
The human gut is inhabited by many organisms, including bacteria and fungi, that may affect human health. However, research on human gut mycobiome is still rare.
