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Archive for the ‘Bone Marrow Stem Cells’ Category

Microplastics dampen the self-renewal of hematopoietic stem cells by disrupting the gut microbiota-hypoxanthine-Wnt … – Nature.com

Mice

C57BL/6J (CD45.2) and C57BL6.SJL (CD45.1) mice were purchased from The Jackson Laboratory and housed under specific pathogen-free conditions. Male and female mice from 8 to 12 weeks were used in experiments and provided with a suitable environment and sufficient water and food. After a week of acclimatization, each mouse was randomly divided into groups, given 100L pure water, 0.01mg/100L, or 0.1mg/100L MPs by oral gavage every two days for five weeks in a gavage experiment (n=5 for each group). For the intravenous injection experiment, MPs were administered into mouse blood via the tail vein at a rate of 0.1g/100L per week for a duration of 4 weeks (n=5 for each group). All animal experiments were first approved by the Laboratory Animal Welfare and Ethics Committee of Zhejiang University (AP CODE: ZJU20220108).

Indocyanine green polystyrene (ICG-PS), polystyrene (PS) and polymethyl methacrylate (PMMA) particles were obtained from Suzhou Mylife Advanced Material Technology Company (China). Polyethylene (PE) particles were purchased from Cospheric (USA). Scanning electron microscopy (SEM, Nova Nano 450, FEI) was used to characterize the primary sizes and shapes of different MPs20. MPs were dispersed in ultrapure water with sonication before dynamic light scattering analysis (Zetasizer, Malvern, UK) to determine the hydrodynamic sizes and zeta potentials49.

Mice were sacrificed and organs were removed within six hours of ICG-PS gavage, including the heart, lung, kidney, spleen, liver, gastrointestinal tissues and bone marrow. Feces were collected 1h before the mice were sacrificed. Both organs and feces were monitored by ex vivo bioluminescence imaging with a small-animal imaging system50 (IVIS Spectrum, PerkinElmer).

For flow cytometry analysis and isolation of hematopoietic stem and progenitor cells, cells were stained with relevant antibodies51 in PBS with 2% fetal bovine serum for 3045min on ice. Antibody clones that were used: Sca-1-PE-Cy7, c-Kit-APC, CD150-PE, CD48-BV421, CD45.1-FITC, CD45.2 PE-Cy5, Gr-1-PE-Cy5, Mac1-PE-Cy5, IgM-PE-Cy5, CD3-PE-Cy5, CD4- PE-Cy5, CD8-PE-Cy5, CD45R-PE-Cy5 and Ter-119-PE-Cy5. Detailed antibody information is summarized in Supplementary Table S6. HSPCs were stained with a lineage antibody cocktail (Gr-1, Mac1, CD3, CD4, CD8, CD45R, TER119 and B220), Sca-1, c-Kit, CD150 and CD48. Cell types were defined as followed: LSK compartment (LinSca-1+c-Kit+), LT-HSC (LSK CD150+CD48), ST-HSC (LSK CD150CD48), MPP2 (LSK CD150+CD48+) and MPP3/4 (LSK CD150CD48+). B cells (CD45.2+Mac1Gr-1+B220+), T cells (CD45.2+Mac1Gr-1+CD3+) and myeloid cells (CD45.2+Mac1+Gr-1). Samples were analyzed on a flow cytometer (CytoFLEX LX, Beckman). For sorting HSCs, lineage antibody cocktail-conjugated paramagnetic microbeads and MACS separation columns (Miltenyi Biotec) were used to enrich Lin cells before sorting. Stained cells were re-suspended in PBS with 2% FBS and sorted directly using the Beckman moflo Astrios EQ (Beckman). Flow cytometry data were analyzed by FlowJo (BD) software.

Apoptosis of cells was detected by Annexin V staining (Yeason, China). After being extracted from the bone marrow of mice, 5106 cells were labeled with different surface markers for 30 to 45min at 4C and then twice rinsed with PBS. Subsequently, the cells were reconstituted in binding buffer and supplemented with Annexin V. After 30min of incubation, flow cytometry was detected in the FITC channel. Cell cycle analysis was performed with the fluorescein Ki-67 set (BD Pharmingen, USA), following the directions provided by the manufacturer. Briefly, a total of 5106 bone marrow cells were labeled with corresponding antibodies, as previously stated. Afterward, the cells were pre-treated with a fixation/permeabilization concentrate (Invitrogen, USA) at 4C overnight and subsequently rinsed with the binding buffer. The cells were stained with Ki-67 antibody for 1h in the dark and then with DAPI (Invitrogen) for another 5min at room temperature. Flow cytometry data were collected by a flow cytometer (CytoFLEX LX, Beckman, USA).

HSCs were sorted by flow cytometry according to the experimental group (ctrl and PSH mice, Rikenellaceae treatment or hypoxanthine treatment). 150 HSCs were seeded in triplicate on methylcellulose media52 (M3434, Stemcell Technologies, Inc.). After 8 days, the number of colonies was counted by microscopy. In addition, 5000 BM cells were seeded and analyzed the same way as HSCs. The cell culture media was diluted in PBS and subjected to centrifugation at 400g for 5min to determine the total cell number.

Recipient mice (CD45.1) were administered drinking water with Baytril (250mg/L) for 7 days pre-transplant and 10 days post-transplant. The day before transplantation, recipients received a lethal dose of radiation (4.5Gy at a time, divided into two times with an interval of 4h). In primary transplantation, 2105 bone marrow cells from the ctrl or PS group (CD45.2) mice and 2105 recipient-type (CD45.1) bone marrow cells were transplanted into recipient mice (CD45.1) mice. Cells were injected into recipients via tail vein injection. Donor chimerism was tracked using peripheral blood cells every 4 weeks for at least 16 weeks after transplantation. For secondary transplantation, donor BM cells were collected from primary transplant recipients sacrificed at 16 weeks after transplantation and transplanted at a dosage of 2106 cells into irradiated secondary recipient mice (9Gy). Analysis of donor chimerism and the cycle of transplantation in secondary transplantation were the same as in primary transplantation.

For limiting dilution assays52, 1104, 5104 and 2105 donor-derived bone marrow cells were collected from ctrl or PS mice (CD45.2) and transplanted into irradiated (9Gy) CD45.1 recipient mice with 2105 recipient-type (CD45.1) bone-marrow cells. Limiting dilution analysis was performed using ELDA software53. 16 weeks after transplantation, recipient mice with more than 1% peripheral-blood multilineage chimerism were defined as positive engraftment. On the other hand, recipient mice undergoing transplantation that had died before 16 weeks post transplantation were likewise evaluated as having failed engraftment54.

For histological analysis, small intestines were collected and fixed in 4% paraformaldehyde and embedded in paraffin, sectioned (5m thickness), and stained with H&E at ZJU Animal Histopathology Core Facility (China). We used Chius scores33,34 to evaluate the damage for each sample. The grade was as follows: 0, normal mucosa; 1, development of subepithelial Gruenhagens space at the tip of villus; 2, extension of the Gruenhagens area with moderate epithelial lifting; 3, large epithelial bulge with a few denuded villi; 4, denuded villi with lamina propria and exposed capillaries; and 5, disintegration of the lamina propria, ulceration, and hemorrhage. For TEM analysis, slices of the small intestine were fixed with 2.5% glutaraldehyde for ultra-microstructure observation of intestinal epithelial cells. The samples were postfixed for one hour at 4C with 1% osmium tetroxide and 30min with 2% uranyl acetate, followed by dehydration with a graded series of alcohol solutions (50%, 70%, 90% and 100% for 15min each) and acetone (100% twice for 20min). Subsequently, they were embedded with epon (Sigma-Aldrich, MO, US) and polymerized. Ultrathin sections (6080nm) were made, and examined using TEM (Tecnai G2 Spirit 120kV, Thermo FEI).

In the short-term and long-term mouse models for MP ingestion, mice were fasted for 4h before oral gavage of FITC-dextran (4kD, Sigma). The fluorescence intensity of FITC-dextran (50mg/100g body weight) was measured in the peripheral blood after 2h of gavage. Fluorescence was measured using a microplate reader (Molecular Devices, SpectraMax iD5) with excitation at 490nm and emission at 520 nm29.

Fecal samples (about 3050mg per sample) were collected from the ctrl, PSL and PSH mice, quickly frozen in liquid nitrogen, and stored at 80C. DNA samples for the microbial community were extracted using E.Z.N.A. Stool DNA Kit (Omega, USA), according to the manufacturers instructions. In brief, polymerase chain reaction (PCR) amplification of prokaryotic 16S rDNA gene V3V4 region was performed using the forward primer 341F (5-CCTACGGGNGGCWGCAG-3) and the reverse primer 805R (5-GACTACHVGGGTATCTAATCC-3)55. After 35 cycles of PCR, sequencing adapters and barcodes were included to facilitate amplification. The PCR products were detected by 1.5% agarose gel electrophoresis and were further purified using AMPure XT beads (Beckman Coulter Genomics, Danvers, MA, USA), while the target fragments were recovered using the AxyPrep PCR Cleanup Kit (Axygen, USA). In addition, the amplicon library was quantified with the Library Quantification Kit for Illumina (Kapa Biosciences, Woburn, MA, USA), and sequenced on the Illumina NovaSeq PE250 platform. In bioinformatics pipeline29,56, the assignment of paired-end reads to samples was determined by their unique barcode, and subsequently shortened by cutting off the barcode and primer sequence. The paired-end reads were combined by FLASH (v1.2.8). Quality filtering on the raw reads was carried out under precise parameters to obtain high-quality clean tags according to fqtrim (v0.94). The chimeric sequences were filtered by Vsearch software (v2.3.4). After the dereplication process using DADA2, we acquired a feature table and feature sequence. The bacterial sequence fragments obtained were grouped into Operational Taxonomic Units (OTUs) and compared to the Greengenes microbial gene database using QIIME2. Alpha diversity and beta diversity were generated by QIIME2, and pictures were drawn by R (v3.2.0). The species annotation sequence alignment was performed by Blast, with the SILVA and NT-16S databases as the alignment references. Additional sequencing results are provided in Supplementary Table S1. The experiment was supported by Lc-Bio Technologies.

The methods for the analysis of feces from HSCT donors were slightly different from those used for mice. All samples were stored in the GUHE Flora Storage buffer (GUHE Laboratories, China). The bacterial genomic DNA was extracted with the GHFDE100 DNA isolation kit (GUHE Laboratories, China) and quantified using a NanoDrop ND-1000 spectrophotometer (Thermo Fisher Scientific, USA). The V4 region of the bacterial 16S rDNA genes was amplified by PCR, with the forward primer 515F (5-GTGCCAGCMGCCGCGGTAA-3) and the reverse primer 806R (5-GGACTACHVGGGTWTCTAAT-3). PCR amplicons were purified with Agencourt AMPure XP Beads (Beckman Coulter, IN) and quantified by the PicoGreen dsDNA Assay Kit (Invitrogen, USA). Following the previously reported steps57, the paired-end 2150bp sequencing was performed on the Illumina NovaSeq6000 platform. The details of bacterial OTUs are summarized in Supplementary Table S5. Sequence data analyses were performed using QIIME2 and R packages (v3.2.0).

For metabolite evaluation, samples from mice feces were prepared and detected as previously described55,58,59. In a nutshell, metabolites were extracted from feces through precooled 50% methanol buffer and stored at 80C before the LCMS analysis. All chromatographic separations were conducted using an ultra-performance liquid chromatography (UPLC) system (SCIEX, UK). A reversed phase separation was performed using an ACQUITY UPLC T3 column (100mm * 2.1mm, 1.8m, Waters, UK). The temperature of the column oven was maintained at 35C and the flow rate was 0.4mL/min. Both positive (the ionspray voltage floating set at 5000V) and negative ion modes (4500V) were analyzed using a TripleTOF 5600 Plus high-resolution tandem mass spectrometer (SCIEX, UK). The mass spectrometry data were obtained in Interactive Disassembler Professional (IDA) mode, with a time-of-flight (TOF) mass range of 60 to 1200Da. The survey scans were acquired in 150 milliseconds and product ion scans with a charge state of 1+ and 100 counts per second (counts/s) were recorded up to 12. Cycle duration was 0.56s. Stringent quality assurance (QA) and quality control (QC) procedures were applied, as the mass accuracy was calibrated every 20 samples and a QC sample was obtained every 10 samples. LCMS raw data files underwent processing in XCMS (Scripps, La Jolla, CA) to perform peak picking, peak alignment, gap filling, and sample normalization. Online KEGG was adopted to annotate metabolites through the matching between the precise molecular mass data (m/z) of samples and those from the database. PCA and volcano plot were utilized to identify ion characteristics that exhibit significant differences between the groups. The details of metabolomes can be found in Supplementary Table S2. The experiment was supported by Lc-Bio Technologies.

Before FMT, SPF mice received a 200L antibiotic treatment (1g/L ampicillin, 0.5g/L neomycin, 0.5g/L vancomycin and 1g/L metronidazole) for three consecutive days by oral gavage. Fresh feces were collected from ctrl or PS mice and resuspended in reduced PBS (0.5g/L cysteine and 0.2g/L Na2S in PBS) at a ratio of about 120mg feces/mL reduced PBS. Feces were then centrifuged at 500g for 1min to remove insolubilize particles25. Recipients (C57BL/6J mice) were administered 100mL of the supernatant from different groups by oral gavage twice every week for 4 weeks. 2 days after the last FMT, recipients were euthanized to analyze the changes in the hematopoietic system.

The Rikenellaceae strain (ATCC BAA-1961), purchased from ATCC, was cultured in an anaerobic chamber using BD Difco Dehydrated Culture Media: Reinforced Clostridial Medium at a temperature of 37C with a gas mixture of 80% N2 and 20% CO2. The final concentration of Rikenellaceae was 2108 viable c.f.u. per 100L and hypoxanthine (200mg/kg, Sigma, Germany) was dissolved in double distilled water29. Mice first received antibiotic treatment (same as FMT) and were then treated by oral gavage with 100L of either Rikenellaceae or hypoxanthine suspension three times a week for 4 weeks. Reinforced Clostridial Medium or double distilled water was used as a vehicle control, respectively. 2 days after the last administration, recipients were euthanized to analyze the changes in the hematopoietic system. To examine the impact of hypoxanthine on HSCs, we exposed bone marrow cells to direct co-culture with hypoxanthine at a concentration of 100pg/mL for a period of 3 days.

Mouse bone marrow cells were harvested by flushing the mices tibia and femur in phosphate buffered saline (PBS) with 2% fetal bovine serum (GIBCO). Harvested cells were grown into 96-well u-bottom plates containing freshly made HSC culture medium (StemSpanTM SFEM, Stemcell Tec.) with SCF (50ng/mL; PeproTech) and TPO (50ng/mL; PeproTech), at 37C with 5% CO2. For HSC culture, the medium was changed every 3 days by manually removing half of the conditioned medium and replacing it with fresh medium60. To assess the effects of WNT10A, IL-17, TNF and NF-kappa B on hematopoiesis, we cultured HSCs in a basic medium and supplemented them with related proteins (10ng/mL; Cosmo Bio, USA) or PBS as a control for two days, followed by flow cytometry analysis. Different concentrations of PS were added to the medium and tested using CCK-8 and FACS to detect the effect of MPs on cultured HSCs.

1104 HSCs were obtained in triplicate from mouse bone marrow cells from the ctrl or PSH group by flow cytometry sorting and RNA was extracted with RNAiso Plus (Takara, Japan) according to the manufacturers protocol. The concentration and integrity of RNA were examined by Qubit 2.0 and Agilent 2100 (Novogene, China), respectively. Oligo (dT)-coated magnetic beads (Novogene, China) were used to enrich eukaryotic mRNA. After cDNA synthesis and PCR amplification, the PCR product was purified using AMPure XP beads (Novogene, China) to obtain the final library. The Illumina high-throughput sequencing platform NovaSeq 6000 was used for sequencing. Analysis of gene expression was calculated by R or the DESeq2 package61. Detailed information regarding RNA-seq is listed in Supplementary Table S3.

For RNA expression analysis, total RNA from bone marrow cells was extracted using Trizol (Invitrogen, US) and resuspended in nuclease-free water. Reverse transcription was performed using the QuantiTect Reverse Transcription kit (Qiagen NV). qPCR was conducted using cDNA, primers and SYBR-green (Takara, Japan) in 20L using the ABI 7500 Q-PCR system62. Results were calculated using the RQ value (RQ=2Ct). Mouse Actin was chosen as the normalization control. Gene-specific primer sequences are shown in Supplementary Table S7.

Bone marrow and Rikenellaceae supernatant in different groups were obtained by centrifugation. Fecal supernatant was obtained from human samples. Hypoxanthine (LANSO, China) and WNT10A (EIAab, China) were measured by ELISA with respective kits according to the manufacturers protocols.

Human feces and peripheral blood samples were obtained from 14 subjects who provided grafts for HSCT patients. They were divided into graft success group and graft failure (GS)/poor graft function (GF/PGF) group, with 7 participants in each group. Research involving humans was approved by the Clinical Research Ethics Committee of the First Affiliated Hospital, College of Medicine, Zhejiang University (IIT20230067B). All participants read and signed the informed consent. Detailed information on patients was listed in Supplementary Table S4.

The Agilent 8700 Laser Direct Infrared Imaging system was utilized for fast and automated analysis of MPs in feces received from donors. An excessive nitric acid concentration (68%) was added to the sample and heated to dissolve the protein. Large particles were first intercepted with a large aperture filter and then filtered by vacuum extraction. After rinsing with ultra-pure water and ethanol several times, the materials, including MPs, were dispersed in the ethanol solution. The LDIR test was carried out when the ethanol was completely volatilized63. The sample of MPs was positioned on the standard sample stage. The stage was then put into the sample stage, and the Agilent Clarity was initiated to advance the sample stage into the sample chamber. The software rapidly scanned the chosen test area using a constant wave number of 1800cm1, and accurately detected and pinpointed the particles within the selected area. The unoccupied area devoid of particles was automatically designated as the background. The background spectrum was gathered and readjusted, followed by the visualization of detected particles and the collection of the whole infrared spectrum. After obtaining the particle spectrum, the spectrum library was utilized to carry out qualitative analysis automatically, including the inclusion picture, size, and area of each particle. The test was supported by Shanghai WEIPU Testing Technology Group.

MPs in peripheral blood from donors were tested by Py-GC/MS. Nitric acid was added to samples for digestion at 110C for 12h, and then used deionized water to make the solution weakly acidic. After concentration, the solution was dribbled into the sampling crucible of Py-GCMS and tested when the solvent in the crucible was completely volatilized17. Various standards of MPs were prepared and analyzed using Py-GCMS in order to construct the quantitative curve. PY-3030D Frontier was employed for lysis, with a lysis temperature set at 550 C. The chromatographic column dimensions were 30m in length, 0.25mm inner diameter, and 0.25m film thickness. The sample was subjected to a heat preservation period of 2min at 40C, followed by a gradual increase in temperature at a rate of around 20C per minute until it reached 320C. The sample was maintained at this temperature for 14min and the entire process takes a total of 30min. The carrier gas utilized was helium, with the ion source temperature of 230C. The split ratio employed was 5:1, and the m/z scan range spanned from 40 to 60064. The experiment was supported by Shanghai WEIPU Testing Technology Group.

Each animal experiment was tested using at least 56 replicates and each in vitro experiment was at least three replicates. Specific replication details are provided in relevant figure captions. Statistical significance was ascertained through unpaired two-tailed t-tests by GraphPad Prism when the P value was less than 0.05. Error bars in all figures indicate the standard deviation (SD).

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Microplastics dampen the self-renewal of hematopoietic stem cells by disrupting the gut microbiota-hypoxanthine-Wnt ... - Nature.com

Hereditary Alzheimer’s Transmitted Via Bone Marrow Transplants – Neuroscience News

Summary: Alzheimers disease, traditionally seen as a brain-centric condition, may have systemic origins and can be accelerated through bone marrow transplants from donors with familial Alzheimers to healthy mice.

A new study underscores the diseases potential transmission via cellular therapies and suggests screening donors for Alzheimers markers to prevent inadvertent disease transfer.

By demonstrating that amyloid proteins from peripheral sources can induce Alzheimers in the central nervous system, this research shifts the understanding of Alzheimers towards a more systemic perspective, highlighting the need for cautious screening in transplants and blood transfusions.

Key Facts:

Source: Cell Press

Familial Alzheimers disease can be transferred via bone marrow transplant, researchers show March 28 in the journalStem Cell Reports. When the team transplanted bone marrow stem cells from mice carrying a hereditary version of Alzheimers disease into normal lab mice, the recipients developed Alzheimers diseaseand at an accelerated rate.

The study highlights the role of amyloid that originates outside of the brain in the development of Alzheimers disease, which changes the paradigm of Alzheimers from being a disease that is exclusively produced in the brain to a more systemic disease.

Based on their findings, the researchers say that donors of blood, tissue, organ, and stem cells should be screened for Alzheimers disease to prevent its inadvertent transfer during blood product transfusions and cellular therapies.

This supports the idea that Alzheimers is a systemic disease where amyloids that are expressed outside of the brain contribute to central nervous system pathology, says senior author and immunologist Wilfred Jefferies, of the University of British Columbia.

As we continue to explore this mechanism, Alzheimers disease may be the tip of the iceberg and we need to have far better controls and screening of the donors used in blood, organ and tissue transplants as well as in the transfers of human derived stem cells or blood products.

To test whether a peripheral source of amyloid could contribute to the development of Alzheimers in the brain, the researchers transplanted bone marrow containing stem cells from mice carrying a familial version of the diseasea variant of the human amyloid precursor protein (APP) gene, which, when cleaved, misfolded and aggregated, forms the amyloid plaques that are a hallmark of Alzheimers disease.

They performed transplants into two different strains of recipient mice: APP-knockout mice that lacked an APP gene altogether, and mice that carried a normal APP gene.

In this model of heritable Alzheimers disease, mice usually begin developing plaques at 9 to 10 months of age, and behavioral signs of cognitive decline begin to appear at 11 to 12 months of age. Surprisingly, the transplant recipients began showing symptoms of cognitive decline much earlierat 6 months post-transplant for the APP-knockout mice and at 9 months for the normal mice.

The fact that we could see significant behavioral differences and cognitive decline in the APP-knockouts at 6 months was surprising but also intriguing because it just showed the appearance of the disease that was being accelerated after being transferred, says first author Chaahat Singh of the University of British Columbia.

In mice, signs of cognitive decline present as an absence of normal fear and a loss of short and long-term memory. Both groups of recipient mice also showed clear molecular and cellular hallmarks of Alzheimers disease, including leaky blood-brain barriers and buildup of amyloid in the brain.

Observing the transfer of disease in APP-knockout mice that lacked an APP gene altogether, the team concluded that the mutated gene in the donor cells can cause the disease and observing that recipient animals that carried a normal APP gene are susceptible to the disease suggests that the disease can be transferred to health individuals.

Because the transplanted stem cells were hematopoietic cells, meaning that they could develop into blood and immune cells but not neurons, the researchers demonstration of amyloid in the brains of APP knockout mice shows definitively that Alzheimers disease can result from amyloid that is produced outside of the central nervous system.

Finally the source of the disease in mice is a human APP gene demonstrating the mutated human gene can transfer the disease in a different species.

In future studies, the researchers plan to test whether transplanting tissues from normal mice to mice with familial Alzheimers could mitigate the disease and to test whether the disease is also transferable via other types of transplants or transfusions and to expand the investigation of the transfer of disease between species.

In this study, we examined bone marrow and stem cells transplantation. However, next it will be important to examine if inadvertent transmission of disease takes place during the application of other forms of cellular therapies, as well as to directly examine the transfer of disease from contaminated sources, independent from cellular mechanisms, says Jefferies.

Funding:

This research was supported by the Canadian Institutes of Health Research, the W. Garfield Weston Foundation/Weston Brain Institute, the Centre for Blood Research, the University of British Columbia, the Austrian Academy of Science, and the Sullivan Urology Foundation at Vancouver General Hospital.

Author: Kristopher Benke Source: Cell Reports Contact: Kristopher Benke Cell Reports Image: The image is credited to Neuroscience News

Original Research: The findings will appear in Stem Cell Reports

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Hereditary Alzheimer's Transmitted Via Bone Marrow Transplants - Neuroscience News

ATG or post-transplant cyclophosphamide to prevent GVHD in matched unrelated stem cell transplantation? | Leukemia – Nature.com

Patient characteristics

The baseline characteristics of the study population are presented in Table1. A total of 8764 patients were included, from which 7725 (88%) received rATG, and 1039 (12%) received PTCy as GVHD prophylaxis.

Overall, the majority of patients were transplanted for acute leukemia (58%), myelodysplastic syndrome (MDS) (19.7%), myeloproliferative neoplasm (MPN) (9.7%) or lymphoma (9%). A high proportion of patients had a low/intermediate Disease Risk Index (DRI, 72.1%), and myeloablative conditioning (MAC) was more frequently performed (53.3%) than reduced intensity conditioning (RIC).

Patients in the rATG group were older, with a median age of 58.6 years (IQR (48.1, 65.4)) vs. 53 years in the PTCy group (IQR 38.6, 62.3) (p<0.01), with a similar proportion of males (57.3% in rATG vs. 58.9% in PTCy, p=0.33), along with a significantly lower use of TBI (14.5% vs. 24.7%, p<0.01) and lower use of MAC (52% vs. 62.3%, p<0.01). Also, the disease relapse index was lower and the year of transplant was more recent in the PTCy group (Table1). The remaining parameters were balanced between the two groups. Median follow up was 2.1 years in both arms. More detailed information is given in Table1.

Univariate outcomes are shown in Figs.1, 2and Table2. The results of the multivariate analyses are summarized in Table3. The P-values and hazard ratios (HR) presented in the following results section are derived from the multivariate analysis.

A NRM; B Overall survival, C Relapse incidence, D Progression-free survival and E GVHD-free relapse-free survival. Cumulative incidences are shown.

A Acute GVHD grades IIIV; B Acute GVHD grades IIIIV, C Chronic GVHD all grades and D Extensive chronic GVHD - Cumulative incidences are shown.

Patients receiving PTCy had a significantly lower NRM as compared to patients receiving rATG (2y incidence: 12.4% vs. 16.1%; HR: 0.72 [95% CI 0.550.94], p=0.016). Similarly, OS and PFS showed a statistically significant and clinically meaningful benefit for PTCy arm, with a higher OS (2y incidence: 73.9% vs. 65.1%; HR: 0.82 [95% CI 0.720.92], p=0.001), and a higher PFS (2y incidence: 64.9% vs. 57.2%; HR: 0.83 [95% CI 0.740.93], p<0.001). RI was lower in the PTCy arm (2y incidence: 22.8% vs. 26.6%; HR: 0.87 [95% CI 0.751.00], p=0.046).

The causes of death are given in Table4. No major differences between the two groups were apparent. Relapse of the underlying malignancy was the most frequent cause of death, accounting for ~50% of total deaths in both arms, followed by NRM causes: infections ~18%, GVHD~16% and other alloSCT-related causes ~8% of total deaths. Secondary malignancies contributed to approximately 1% of total deaths.

Overall chronic GVHD was lower in the PTCy group (2y incidence: PTCy 28.4% vs. rATG 31.4%; HR: 0.77 [95% CI 0.630.95], p=0.012). Extensive chronic GVHD was also reduced in patients receiving PTCy vs. rATG: (2y incidence: 11.9% vs. 13.5%; HR: 0.75 [95% CI 0.620.91], p=0.004).

The incidence of acute GVHD grades II-IV in patients receiving PTCy, compared to those receiving ATG was not statistically significant: (100d incidence: 24.1% vs. 26.5%; HR: 0.85 [95% CI 0.691.04], p=0.11). Similarly, for severe acute GVHD grades IIIIV (100d incidence: 8.7% vs. 9.7%; HR: 0.76 [95% CI 0.551.05], p=0.091).

GRFS was significantly higher in the PTCy arm compared to the rATG arm (2y incidence: 51% vs. 45%; HR: 0.86 [95% CI 0.750.99], p=0.035).

The EBMT Database does not contain data on graft failure/rejection. To get insight into the initial grafts success and any subsequent requirement for additional transplantation procedures, we investigated neutrophil recovery after the first alloSCT as well as the incidence of a second alloSCT. The median incidence of neutrophil recovery at days +30 and +60 in the ATG vs. PTCy groups was: d+30 ATG 96% (IC95% 95.596.4) vs. PTCy 91% (8992.7) and d+60 ATG 97.9% (97.698.2) vs. PTCy 97.4% (96.298.3). The median incidence of a second alloSCT at 2 years was 4.3% (3.84.8) in the ATG group and 3.2% (2.24.6) in the PTCy group.

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ATG or post-transplant cyclophosphamide to prevent GVHD in matched unrelated stem cell transplantation? | Leukemia - Nature.com

New Allo-HCT Approach Boosts Immune Response, Survival – Targeted Oncology

While ex vivo CD34-selected allogeneic hematopoietic stem cell transplants (HCTs) are promising treatments for patients with hematologic and myeloid malignancies, they can be limited by delayed immune recovery and increased risk of death not caused by relapse.

A late-breaking abstract presented at the 2024 Transplantation and Cellular Therapy Tandem Meetings investigated a new approach to allogeneic HCT. Investigators of the phase 2 PRAISE-IR study (NCT04872595) explored using a model-based approach to determine the optimal dose of antithymocyte globulin (ATG), which is used to prevent graft-vs-host disease after transplant. Previous studies suggested high ATG exposure might contribute to nonrelapse mortality.

According to Michael Scordo, MD, the model successfully achieved a low posttransplant ATG exposure, and immune reconstitution by day 100 was achieved in 69% of patients, meeting the studys primary end point. Further, the 2-year rates of nonrelapse mortality and relapse were 9% and 13%, respectively, and relapse-free survival and overall survival rates were high at 78% and 86%, respectively.

These findings suggest that using a model to determine the ATG dose for ex vivo CD34-selected allogeneic HCT can lead to improved immune reconstitution and excellent survival outcomes. This approach may help reduce nonrelapse mortality previously observed in other trials and improve the safety and effectiveness of this type of transplant.

In an interview with Targeted OncologyTM, Scordo, bone marrow transplant specialist and cellular therapist at Memorial Sloan Kettering Cancer Center in New York, New York, discussed the findings from this study and their implications for the allogeneic HCT treatment landscape.

Targeted Oncology: What was the rationale or inspiration for the study you presented at the Tandem Meetings?

Scordo: Ex vivo CD34-selected [allogeneic] transplant is one of the many methods of reducing graft-vs-host disease. It uses a myeloablative conditioning platform and integrates ATG, antithymocyte globulin, into that platform to help reduce the risk of rejection. This has been well studied over the years, but 1 of the downsides of this approach is the delayed immune recovery, particularly the T-cell immune recovery that occurs after [allogeneic] transplant with this approach. What we did based on a recent publication that we have from last year was we used a different dosing of ATG to ensure that the T-cell immune recovery after [allogeneic] transplant using ex vivo CD34 selection would be improved.

What are some of the unmet needs in this space?

There are many methods to reduce graft-vs-host disease after transplant CD34 selection. Many of the other methods including posttransplant cyclophosphamide [PTCy], which has now become a standard of care, are out there and should be used in the appropriate setting. In matched donor transplants, ex vivo CD34 selection is one of the methods of being able to use an ablative or intensive conditioning regimen with very low rates of particularly chronic graft-vs-host disease. We saw this as an opportunity to improve on this platform significantly, using a novel approach but a simple approach.

What were the goals of this study?

The primary end point of the study was the ability to improve the CD34 T-cell immune recovery by day 100 after transplant. This was a sort of a validated predictor in other studies. We had key secondary end points that included nonrelapse mortality, relapse rates, progression-free, and overall survival. With the primary end point, we exceeded that end point. With our trial, about 70% of our 56 patients achieved this appropriate immune recovery by day 100, which was significantly higher than our historical numbers had shown.

What were some of the other findings?

Aside from achieving the primary end point, we saw very low rates of nonrelapse mortality at 2 years, estimated at 8%, which is much lower than some of the previously published data using this platform in the last couple of years. [We also saw] low relapse rates [of] about 12% at 2 years and very favorable progression-free and overall survival, which was 80% and 87%, respectively, at 2 years.

What are some of the takeaways?

I look at this as a simple but novel approach to improving on a platform. We have existing platforms that work well, but we can improve them doing well. To community oncologists, I would say that for patients with myeloid malignancies, there are many different types of transplants that can be done safely and effectively. The appropriate choice of a platform really depends on many factors. We can improve on all these platforms individually, including PTCy. [For] ex vivo CD34 selection, I look at this as a method of just improving on what we have already shown to be an effective platform, being able to use dose-intensive chemotherapy or total body radiation to achieve maximal disease control but making the platform safe and tolerable.

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New Allo-HCT Approach Boosts Immune Response, Survival - Targeted Oncology

New immunotherapy could make blood more ‘youthful,’ mouse study hints – Livescience.com

Scientists reversed some signs of immune aging in mice with a new treatment that could one day potentially be used in humans.

The new immunotherapy works by disrupting a natural process by which the immune system becomes biased towards making one type of cell as it ages.

The mouse study is an "important" proof-of-concept, but it's currently difficult to gauge the significance of the findings, Dr. Janko . Nikolich-Zugich, a professor of immunobiology at the University of Arizona who was not involved in the research, told Live Science in an email. More work is needed to see how well the therapy shifts the immune system into a more youthful, effective state.

All blood cells, including immune cells and the red blood cells that carry oxygen around the body, start life as hematopoietic stem cells (HSC) in the blood and bone marrow, the spongy tissue found within certain bones. HSCs fall into two main categories: those destined to become so-called myeloid cells and those that will develop into lymphoid cells.

Myeloid cells include red blood cells and immune cells belonging to our broadly reactive first line of defense against pathogens, including cells called macrophages that trigger inflammation. Lymphoid cells include cells that develop a memory of germs, such as T and B cells.

Related: 'If you don't have inflammation, then you'll die': How scientists are reprogramming the body's natural superpower

As we age, the HSCs slated to become myeloid cells gradually increase in number and eventually outnumber the lymphoid stem cells. This means we can't respond to infections as well when we're older as when we're young, and we're more likely to experience chronic inflammation triggered by increasing levels of myeloid cells that trigger inflammation.

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In the new study, published Wednesday (March 27) in the journal Nature, scientists developed an antibody-based therapy that selectively targets and destroys the myeloid HSCs, thus restoring the balance of the two cell types and making the blood more "youthful." The antibodies latch onto the targeted cells and flag them to be destroyed by the immune system.

The authors injected the therapy into mice aged 18 to 24 months, or roughly the equivalent of being between 56 and 69 years old as a human.

They then extracted HSCs from the mice after treatment and analyzed them, revealing the rodents had a smaller percentage of the myeloid HSCs than untreated mice of the same age.

This effect lasted for two months. Compared with untreated mice, the treated mice also produced more naive T cells and mature B cells. These cells can go on to form memory cells, which are directly involved in the immune attack; in the case of the B cells, they can form antibody-producing plasma cells.

"Not only did we see a shift toward cells involved in adaptive immunity, but we also observed a dampening in the levels of inflammatory proteins in the treated animals," Dr. Jason Ross, lead study author and postdoctoral researcher at Stanford University, said in a statement. Specifically, the researchers saw that the levels of one proinflammatory protein fell in the treated mice. This protein, called IL-1beta, is mainly made by myeloid cells.

Eight weeks post-treatment, the researchers vaccinated the mice against a virus they'd never been exposed to before. The mice that had received the immunotherapy had more apt immune responses to vaccination than the untreated mice, producing more T cells against the germ.

"We believe that this study represents the first steps in applying this strategy in humans," Ross said. However, other experts have cautioned against jumping to conclusions.

Nikolich-Zugich noted that, although the researchers measured changes in the numbers of naive T cells in the mice, they didn't look at the function of the organ that makes them: the thymus. The team also saw reductions only in IL-1beta and not other inflammatory proteins. They also didn't test whether the mice's baseline immunity to new infections could be improved with this therapy, without vaccination, he said.

Furthermore, the study didn't consider potential long-term side effects of the treatment, such as anemia, or a deficiency in red blood cells, said Dr. Ilaria Bellantuono, a professor in musculoskeletal aging at the University of Sheffield in the U.K. who was not involved in the research.

Although an "interesting" study, more work is needed to understand whether it can bring "meaningful changes" in the immune system, Bellantuono told Live Science in an email, whether that of mice or humans.

Ever wonder why some people build muscle more easily than others or why freckles come out in the sun? Send us your questions about how the human body works to community@livescience.com with the subject line "Health Desk Q," and you may see your question answered on the website!

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New immunotherapy could make blood more 'youthful,' mouse study hints - Livescience.com

Allogeneic hematopoietic stem-cell transplantation for patients with Richter transformation: a retrospective study on … – Nature.com

Allogeneic hematopoietic stem-cell transplantation for patients with Richter transformation: a retrospective study on ...  Nature.com

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Allogeneic hematopoietic stem-cell transplantation for patients with Richter transformation: a retrospective study on ... - Nature.com

Long term outcomes of nonmyeloablative allogeneic stem cell transplantation with TSEB TLI and ATG for Mycosis … – Nature.com

Long term outcomes of nonmyeloablative allogeneic stem cell transplantation with TSEB TLI and ATG for Mycosis ...  Nature.com

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Long term outcomes of nonmyeloablative allogeneic stem cell transplantation with TSEB TLI and ATG for Mycosis ... - Nature.com

BioRestorative Therapies to Participate in the 36th Annual ROTH Conference – Yahoo Finance

BioRestorative Therapies to Participate in the 36th Annual ROTH Conference  Yahoo Finance

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BioRestorative Therapies to Participate in the 36th Annual ROTH Conference - Yahoo Finance

Iron restriction keeps blood stem cells young, researchers find – Phys.org

Iron restriction keeps blood stem cells young, researchers find  Phys.org

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Iron restriction keeps blood stem cells young, researchers find - Phys.org

Effective treatment of Clostridioides difficile infection improves survival and affects graft-versus-host disease: a … – Nature.com

Effective treatment of Clostridioides difficile infection improves survival and affects graft-versus-host disease: a ...  Nature.com

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Effective treatment of Clostridioides difficile infection improves survival and affects graft-versus-host disease: a ... - Nature.com

Woman, 22, With Leukemia Recalls Symptoms And New Treatment She Received: EXCLUSIVE – TODAY

Woman, 22, With Leukemia Recalls Symptoms And New Treatment She Received: EXCLUSIVE  TODAY

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Woman, 22, With Leukemia Recalls Symptoms And New Treatment She Received: EXCLUSIVE - TODAY

This Swedish startup wants to reduce the cost, and controversy, around stem cell production – TechCrunch

This Swedish startup wants to reduce the cost, and controversy, around stem cell production  TechCrunch

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This Swedish startup wants to reduce the cost, and controversy, around stem cell production - TechCrunch

How an MS friendship led to HSCT and a love of running – Multiple Sclerosis News Today

How an MS friendship led to HSCT and a love of running  Multiple Sclerosis News Today

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How an MS friendship led to HSCT and a love of running - Multiple Sclerosis News Today

Team demonstrates fabrication method to construct 3D structures that mimic bone microstructure – Phys.org

Team demonstrates fabrication method to construct 3D structures that mimic bone microstructure  Phys.org

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Team demonstrates fabrication method to construct 3D structures that mimic bone microstructure - Phys.org

Stem Cell Therapy for Crohns Disease Shows Promising Results – RegMedNet

Stem Cell Therapy for Crohns Disease Shows Promising Results  RegMedNet

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Stem Cell Therapy for Crohns Disease Shows Promising Results - RegMedNet

A SMART way to efficiency and cost reduction in stem cell manufacturing – pharmaphorum

A SMART way to efficiency and cost reduction in stem cell manufacturing  pharmaphorum

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A SMART way to efficiency and cost reduction in stem cell manufacturing - pharmaphorum

UAE scientists hold hope of finding MS cure as patient walks again after treatment – The National

UAE scientists hold hope of finding MS cure as patient walks again after treatment  The National

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UAE scientists hold hope of finding MS cure as patient walks again after treatment - The National

Analyzing the Impact of Eltrombopag on Platelet Engraftment Following Haploidentical Bone Marrow Transplantation – Physician’s Weekly

Analyzing the Impact of Eltrombopag on Platelet Engraftment Following Haploidentical Bone Marrow Transplantation  Physician's Weekly

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Analyzing the Impact of Eltrombopag on Platelet Engraftment Following Haploidentical Bone Marrow Transplantation - Physician's Weekly

Young cancer patient with ‘no hope’ of recovery saved by baby’s umbilical cord – Study Finds

Young cancer patient with 'no hope' of recovery saved by baby's umbilical cord  Study Finds

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Young cancer patient with 'no hope' of recovery saved by baby's umbilical cord - Study Finds

Second haploidentical bone marrow transplantation with antithymocyte antibody-containing conditioning regimen for … – Nature.com

Second haploidentical bone marrow transplantation with antithymocyte antibody-containing conditioning regimen for ...  Nature.com

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Second haploidentical bone marrow transplantation with antithymocyte antibody-containing conditioning regimen for ... - Nature.com

Global Stem Cell Therapy Industry Outlook to 2028, Driven by Therapeutic Innovations and Clinical Advancements … – Yahoo Finance

Global Stem Cell Therapy Industry Outlook to 2028, Driven by Therapeutic Innovations and Clinical Advancements ...  Yahoo Finance

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Global Stem Cell Therapy Industry Outlook to 2028, Driven by Therapeutic Innovations and Clinical Advancements ... - Yahoo Finance

Stem cell study shows how gene activity modulates the amount of immune cell production in mice – Medical Xpress

Stem cell study shows how gene activity modulates the amount of immune cell production in mice  Medical Xpress

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Stem cell study shows how gene activity modulates the amount of immune cell production in mice - Medical Xpress

Reconstructing family trees of blood cells may help predict disease – STAT – STAT

Reconstructing family trees of blood cells may help predict disease - STAT  STAT

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Reconstructing family trees of blood cells may help predict disease - STAT - STAT

Dr Aldoss on the Use of Revumenib in KMT2A-Rearranged Acute Leukemia – OncLive

Dr Aldoss on the Use of Revumenib in KMT2A-Rearranged Acute Leukemia  OncLive

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Dr Aldoss on the Use of Revumenib in KMT2A-Rearranged Acute Leukemia - OncLive

Donating Bone Marrow and Stem Cells: The Process and What To Expect – On Cancer – Memorial Sloan Kettering

Donating Bone Marrow and Stem Cells: The Process and What To Expect  On Cancer - Memorial Sloan Kettering

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Donating Bone Marrow and Stem Cells: The Process and What To Expect - On Cancer - Memorial Sloan Kettering

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