Outcome Of Transplantation Research Articles

Retinal Ganglion Cell Transplantation: Differentiation and Quantification Strategies.

Glaucoma is a leading cause of irreversible blindness worldwide. Current treatments focus on reducing intraocular pressure but cannot restore lost visual function once it is lost due to retinal ganglion cell (RGC) degeneration and death. Recent advances suggest that transplantation of stem cell-derived RGCs could offer new therapeutic approaches for glaucoma and vision restoration. Here, we present a detailed protocol for differentiating human RGCs from embryonic stem cells using both three-dimensional retinal organoid and two-dimensional culture approaches. Following differentiation, we describe methods for isolating and purifying retinal ganglion cells from these cultures and their subsequent transplantation into the mouse retina, with careful monitoring and postoperative care to ensure successful integration. Finally, we describe a quantitative method for assessing transplantation outcomes involving confocal imaging of retinal flat-mounts and custom ImageJ and MATLAB scripts to map and analyze the spatial distribution of donor RGCs within the host retina. Altogether, this approach provides a robust framework for exploring RGC transplantation as a potential therapy for vision loss in glaucoma and other optic neuropathies.

A machine learning-based workflow for predicting transplant outcomes in patients with sickle cell disease.

Allogeneic haematopoietic cell transplantation (HCT) with HLA-matched sibling donor remains the most established curative therapeutic option for patients with sickle cell disease (SCD). However, it is not without risks, highlighting the need for a risk stratification system. Utilizing a machine learning (ML) approach that combines clinical and imaging variables, we identified red cell distribution width and renal organ damage as important risk factors for patients undergoing HCT. This ML-based algorithm, similar to an approach previously reported for predicting mortality in patients with SCD, should be applicable to risk factor discovery in similar studies.

A systematic review on the epidemiology and treatment options of multiple Myeloma in Asia

Multiple myeloma (MM) accounts for almost 15 % of all neoplastic malignancies around the globe. This systematic review intends to analyse data on the treatment and management of MM in selected regions in Asia to identify and prioritize areas that need attention. A comprehensive review of original articles, published in English from 2005 to 2022, derived from the PubMed/MEDLINE database was conducted based on the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. There were 98 studies from select regions of Asia (China, India, Taiwan, Hong Kong, and Singapore) on newly diagnosed MM and relapsed/refractory MM. This review evaluated the trends in disease outcomes with the gradual shift in treatment regimens from doublet to triplet. Additionally, this review also explored autologous stem cell transplant outcome and anti-B-cell maturation antigen (BCMA) chimeric antigen receptor (CAR) T-cell therapy in MM patients. This is the first systematic review attempting to collect data on the utility and comparison of innovative agents and modifications in treatment regimens in the context of the Asian population. This review established that the body of evidence for the management of MM was generally of poor quality and there is a need for more versatile studies in the region. Novel and innovative drug regimens may help in combating the illness but consorted efforts by researchers, industry partners, policymakers, and the government are key factors in the long-term survival of MM patients. In the current systematic review, the authors have tried to give a comprehensive account of the available treatments, trends in MM management and prognosis for MM in Asia.

Comparative outcomes of various transplantation platforms, highlighting haploidentical transplants with post-transplantation cyclophosphamide for adult T-cell leukaemia/lymphoma.

This study retrospectively compared outcomes of various allogeneic haematopoietic cell transplantation (allo-HCT) platforms in patients with adult T-cell leukaemia/lymphoma. Platforms included human leukocyte antigen (HLA)-haploidentical-related donors using post-transplant cyclophosphamide (PTCY), HLA-matched related donors (MRD), HLA-matched unrelated donors (MUD) and cord blood transplantation (CBT). Patients who underwent their first allo-HCT between 2016 and 2021 were included. Outcomes analysed were overall survival (OS), relapse and non-relapse mortality (NRM). Seven hundred patients were included (PTCY, n = 121; MRD, n = 91; MUD, n = 160; CBT, n = 328). With a median follow-up of 794 days for survivors, 2-year OS was 48.1% (PTCY), 48.8% (MRD), 48.4% (MUD) and 34.6% (CBT); the respective 2-year cumulative incidence of relapse was 37.1%, 47.5%, 33.9% and 45.1% and that of NRM was 24.2%, 19.8%, 24.7% and 27.3%. PTCY was associated with delayed platelet engraftment relative to MRD and MUD. There was no increase in the incidence of severe acute or chronic graft-versus-host disease. In the PTCY group, poor performance status was a significant predictor of inferior OS, and infused CD34+ cell numbers of less than 5 × 106/kg were associated with delayed neutrophil and platelet engraftment. These results suggest that allo-HCT with PTCY is a safe and effective platform for patients with adult T-cell leukaemia/lymphoma.

Revisiting the Prognostic Influences of Donor-Recipient Size Mismatch in Deceased Donor Liver Transplantation.

Liver transplantation (LT) outcomes are influenced by donor-recipient size mismatch. This study re-evaluated the impact on graft size discrepancies on survival outcomes. Data from 53 389 adult LT recipients from the United Network for Organ Sharing database (2013-2022) were reviewed. The study population was divided by the body surface area index (BSAi), defined as the ratio of donor body surface area (BSA) to recipient BSA, into small-for-size (BSAi < 0.78), normal-for-size (BSAi 0.78-1.24), and large-for-size (BSAi > 1.24) grafts in deceased donor LT (SFSD, NFSD, and LFSD). Multivariate Cox regression and Kaplan-Meier survival analyses were conducted. The frequency of size mismatch in deceased donor LT increased over the past 10 y. SFSD had significantly worse 90-d graft survival (P < 0.01), and LFSD had inferior 1-y graft survival among 90-d survivors (P = 0.01). SFSD was hazardous within 90 d post-LT because of vascular complications. Beyond 1 y, graft size did not affect graft survival. LFSD risk within the first year was mitigated with lower model for end-stage liver disease (MELD) 3.0 scores (<35) or shorter cold ischemia time (<8 h). The negative impacts on donor-recipient size mismatch on survival outcomes are confined to the first year post-LT. SFSD is associated with a slight decrease in 90-d survival rates. LFSD should be utilized more frequently by minimizing cold ischemia time to <8 h, particularly in patients with MELD 3.0 scores below 35. These findings could improve donor-recipient matching and enhance LT outcomes.

Outcome of Allogeneic Hematopoietic Stem Cell Transplantation in Adult Patients with Acute Lymphoblastic Leukemia: Results of a Single-Center Study.

Despite the adoption of pediatric-like chemotherapy protocols, the introduction of new immunotherapies and a better understanding of the oncogenic landscape, the outcome for adult patients with acute lymphoblastic leukemia (ALL) remain substantially dismal. The aim of the present study was to evaluate the outcome in terms of survival in a cohort of adult patients with ALL who received allogeneic hematopoietic stem cell transplantation (alloSCT) between 2013 and 2023. This was a single-center observational retrospective study including all consecutive adult patients with ALL who received an alloSCT between April 2013 and April 2023 at the Stem Cell Transplant Center AOU Città della Salute e della Scienza of Torino. The primary endpoints were overall survival (OS), graft-versus-host disease (GVHD) Relapse-Free Survival (GRFS), Leukemia-Free Survival (LFS) and cumulative incidence (CI) of Non-Relapse Mortality (NRM). The 4-year OS and LFS were 63.4% and 48.1%, respectively, and the 1-year GRFS was 42.9%. The 1-year CI of bloodstream infections (BSI), invasive fungal infections and NRM were 38%, 7% and 18.4%, respectively. Multivariate analysis showed that the use of total body irradiation (TBI), a time interval from diagnosis to alloSCT 7 months and female gender were factors significantly associated with better OS. Relapse of the underlying malignancy and BSI were the main causes of death. Our study suggests that alloSCT from a matched sibling donor (MSD) and alternative donors may be considered an effective tool for patients with ALL achieving a CR.

Prognostic significance of preoperative nutritional status for heart transplantation patients

BackgroundThe association between malnutrition and outcomes of heart transplantation (HTx) has not been well studied. The purpose of this article was to evaluate the prognostic value of three different nutrition indices in HTx, including CONUT (Controlling Nutritional Status), NRI (Nutritional Risk Index) and GNRI (Geriatric Nutritional Risk Index).MethodsA total of 438 patients who underwent THx from January 2015 to December 2020 were included in this study. The nutritional status of the patients was evaluated by CONUT, NRI and GNRI. Kaplan-Meier (KM) curves were constructed to compare the difference in overall survival (OS) between the normal and malnutrition groups in each index. Cox regression analysis was used to identify the independent risk factors of OS. The predictive power was compared by time-dependent ROC and time-dependent ccurves. Logistic regression model was used to evaluate the relationship between these three nutrition indices and postoperative clinical events.Results336 (76.7%), 183 (43.8%), and 190 (43.4%) patients had malnutrition according to CONUT, NRI and GNRI calculations. 102 (23.3%) patients had died at the end of follow-up. After adjustment for confounding variables, multivariate Cox analysis showed that CONUT [HR 1.286 (95%CI 1.166 ~ 1.419); p < 0.001], NRI [HR 0.942 (95%CI 0.923 ~ 0.962); p < 0.001] and GNRI [HR 0.959 (95%CI 0.939 ~ 0.979); p < 0.001] were all independent predictors for OS. The predictive power of CONUT score was higher than that of NRI (p = 0.045) and GNRI (p < 0.001). Regarding the postoperative complications, multivariate logistic regression model showed that malnutrition assessed by CONUT [HR 1.156 (95%CI 1.032 ~ 1.294); p = 0.012] and NRI [HR 1.543 (95%CI 1.008 ~ 2.362); p = 0.046] was independent risk factors for posttransplant infections.ConclusionPoor nutritional status, as assessed by CONUT, NRI and GNRI, was associated with an increased risk of mortality after HTx. CONUT displayed the highest predictive power compared to the other two indices. CONUT and NRI were also independently associated with posttransplant infections.

Improved Outcomes of Liver Transplantation in Patients With Hepatitis C, Following the Introduction of Innovative Antiviral Therapies

BackgroundThe treatment landscape for Hepatitis C virus (HCV) underwent a significant shift with the introduction of direct-acting antiviral (DAA) medications in late 2013. This study aimed to evaluate the impact of DAAs on liver transplantation outcomes, examining both the benefits and any potential drawbacks associated with their use. Methods and materialsA retrospective registry analysis of the United Network for Organ Sharing database was done for liver transplants in patients diagnosed with Hepatitis C, that were performed in the United States from January 2000 to May 2020. ResultsThe study was divided into 2 subgroups, based on the timing of new DAA medication FDA approved. The only significant difference between the two cohorts is the recipient’s age. The data analysis showed a significant overall 5-year graft survival improvement in the 2014-2020 group compared to the 2000-2013 group, from a mean of 64.8% in 2000-2013 to a mean of 76% in 2014-2020 (P<0.001). Interestingly, when we compared the 5-year graft survivals with recipients who had a donor above age 50, the graft survival rate difference was even more significant (74% vs. 56%, P<0.001), as some studies have shown a suboptimal graft outcome, when the donor age is above 40 years old. Not only has the utilization of DCD livers increased significantly after 2014, but the graft survival in this cohort has also been significantly higher.(P<0.001). ConclusionThe emergence of DAAs in 2013 marked a watershed moment in the management of HCV, offering high cure rates, minimal side effects, and shorter treatment durations, to a point that the short- and long-term outcomes of liver transplant for HCV is almost equal to the other causes of liver transplantation.

Impact of Pelvic Calcification Severity on Renal Transplant Outcomes: A Prospective Single-Center Study.

Background: Vascular calcifications (VC) are increasingly prevalent in patients with chronic kidney disease. This study aimed to assess the incidence of iliac artery calcifications in kidney transplant (KT) patients and explore the relationship between iliac VC burden measured by pelvic calcification score (PCS) and renal transplant outcomes. Methods: This prospective study involved 79 KT recipients. VC quantification, using a pre-transplant computed tomography (CT) scan, was performed by assessing calcifications in the common and external iliac arteries bilaterally, resulting in an overall PCS ranging from 0 (no calcifications) to 44 (extensive calcifications). Based on PCS values, patients were divided into three equal-sized groups: PCS Group 1 (PCS 0-4), PCS Group 2 (PCS 5-19), and PCS Group 3 (PCS > 19). Post-transplant outcomes tracked for at least 1 year were patient and graft survival, graft function (urea, creatinine, MAG-3 clearance), and incidence of MACE during the first post-transplant year. Results: Calcifications were present in at least one arterial segment in 61 patients (77.2%). One-year patient survival was 95%, and one-year graft survival was 92.4%. Patients in PCS Group 3 had significantly lower one-year patient and graft survival compared to those in PCS Group 1 and 2 (p = 0.006 and p = 0.008, respectively). MACE and renal function indicators 1-year post-transplant were similar across all PCS groups. Conclusions: Our study demonstrated that a significant majority of KT recipients exhibited iliac VC during pre-transplant CT assessments. Patients in PCS Group 3 exhibited significantly lower one-year patient and graft survival rates compared to those in PCS Groups 1 and 2, indicating that this subgroup may require more intensive post-transplant monitoring and management.

Outcomes of Multiorgan Heart Transplant Between Donation After Circulatory Death and Brain Death.

There is insufficient data on the outcomes of donation after circulatory death (DCD) multiorgan transplant that includes heart. The primary objective of this study is to compare the overall survival outcomes of DCD and donation after brain death (DBD) multiorgan transplants. We identified all heart transplant patients from 2019 to June of 2023 using the United Network for Organ Sharing (UNOS) Database who also received an additional organ (kidney, liver, and lungs). A total of 1,844 DBD and 91 DCD multiorgan transplants occurred within the study period, the majority being combined heart-kidney transplantation. More patients were listed at a higher status in the DBD group (p < 0.05) and were in the intensive care unit (ICU) before transplant (p < 0.05). Despite the higher ischemia time in the DCD group (p < 0.05), the overall unmatched survival did not differ between the two groups (p < 0.05). Within the heart-kidney transplants, the overall survival between DBD and DCD heart-kidney transplants did not differ in either unmatched or matched groups (unmatched p = 0.5, matched p = 0.5). In conclusion, the data on the outcomes of DCD multiorgan transplants are limited. Still, our analysis of the currently available data suggests that the overall survival is comparable in the DCD multiorgan transplants.

Contribution of tryptophan and its metabolites to transplant outcome: a mini-review.

Long-term tolerance in the absence of immunosuppressive drugs is a major goal in the transplantation field, not yet attained. Recent research on the role of commensal microbiota in the control of immunity has opened new avenues for the search of novel clinical interventions. Indeed, products of intestinal metabolism generated by both host cells and the microbiota have been identified as modulators of the immune response. Among these, tryptophan (Trp) and its derivatives are being investigated to understand their impact on alloimmunity and their potential usefulness as therapeutic targets to improve allograft survival. Here, we reviewed the latest findings on the contribution of Trp metabolic pathways to transplant outcomes.

Quantifying measurement uncertainty in renal transplant biopsy assessment.

Renal transplant biopsies provide insights into graft health and support decision making. The current evidence on links between biopsy scores and transplant outcomes suggests there may be numerous factors affecting biopsy scores. Here we adopt measurement science approach to investigate the sources of uncertainty in biopsy assessment and suggest techniques to improve its robustness. Histological assessments, Remuzzi scores, biopsy processing and clinical variables are obtained from 144 repeat biopsies originating from 16 deceased-donor kidneys. We conducted sensitivity analysis to find the morphometric features with highest discriminating power and studied the dependencies of these features on biopsy and stain type. The analysis results formed a basis for recommendations on reducing the assessment variability. Most morphometric variables are influenced by the biopsy and stain types. The variables with the highest discriminatory power are sclerotic glomeruli counts, healthy glomeruli counts per unit area, percentages of interstitial fibrosis and tubular atrophy as well as diameter and lumen of the worst artery. A revised glomeruli adequacy score is proposed to improve the robustness of the glomeruli statistics, whereby a minimum of 104 µm2 of cortex tissue is recommended to keep type 1 and type 2 error probabilities below 0.15 and 0.2. The findings are transferable to several biopsy scoring systems. We hope that this work will help practitioners to understand the sources of statistical uncertainty and improve the utility of renal biopsy.

Real World Predictors, Timing, and Outcomes of Autologous Stem Cell Transplantation in Patients with Multiple Myeloma

Real World Predictors, Timing, and Outcomes of Autologous Stem Cell Transplantation in Patients with Multiple Myeloma

Navigating 'grey areas' and challenges during evaluation of transplant eligibility in specific myelofibrosis populations: a perspective on behalf of the Chronic Malignancies Working Party of the EBMT.

Significant efforts have been made to effectively select myelofibrosis (MF) patients who can benefit from allogeneic hematopoietic cell transplantation (allo-HCT), the only current cure for MF. The recent EBMT/ELN 2024 recommendations offer valuable guidance for hematologists and transplant physicians. However, several grey areas remain in day-to-day clinical practice regarding the feasibility and optimal preparation for transplantation in patients with this disease. Effective spleen size reduction, often achieved with JAK inhibitors, appears crucial for transplant success. For resistant cases, switching JAK inhibitors, splenectomy, or spleen irradiation may be considered, taking into account patient profiles, treatment availability and center preferences. Managing splanchnic vein thromboses, portal, and pulmonary hypertension is critical as these conditions may affect transplant outcomes. Cytopenias, particularly transfusion-dependent anemia and thrombocytopenia, complicate treatment and impact on outcomes, though new drugs show promise. Comorbidities play a significant role and tools like the Hematopoietic Cell Transplantation-Comorbidity Index (HCT-CI) and frailty assessments are useful for evaluating transplant risks while allowing the implementation of corrective measures. Especially in low- and medium-income countries where access to novel therapies may be challenging, allo-HCT still represents an attractive therapeutic option for MF. Future directions include integrating new therapeutics into the transplant algorithm and leveraging artificial intelligence for more informed risk assessment, highlighting the need for tailored approaches to improve allo-HCT outcomes in such a setting.

Where your liver lives matters: How neighborhoods shape transplant outcomes.

Where your liver lives matters: How neighborhoods shape transplant outcomes.

Extracorporeal membrane oxygenation ameliorate hepatic injury in brain death rat donors with hemodynamic instability.

Donation after brain death (DBD) serves as the primary source for liver transplantation. However, livers obtained through DBD often incur damage due to unstable hemodynamics, potentially impacting transplantation outcomes. Extracorporeal Membrane Oxygenation (ECMO) emerges as an optimal technique for donor liver retrieval and has found application in clinical settings. Despite its clinical implementation, the precise mechanisms through which ECMO enhances liver functions remain elusive. This study aims to investigate the mechanisms underlying how ECMO ameliorates liver function in brain-dead donors. We randomly assigned 18 male Sprague-Dawley (SD) rats (350 ± 50 g) into three groups: Con (n = 6), DBD-assisted drug (n = 6), and DBD-assisted ECMO (n = 6). After 3 h of ECMO, the rats were sacrificed. We assessed and compared changes in heart rate, blood pressure, cumulative liver damage (evaluated through HE and TUNEL staining), serum levels of AST and ALT, alterations in serum oxidative stress factors (MDA, H2O2, SOD, and 8-OHdG), and serum concentrations of related inflammatory factors (interleukin [IL]-1β, IL-6, IL-8, and TNF-α) among rats in the Con, DBD-assisted drug, and DBD-assisted ECMO groups. Subsequently, we established a rat orthotopic liver transplantation (OLT) model and transplanted livers obtained through the aforementioned methods. The post-transplantation status of the livers was observed. After 3 h of brain death, liver injury worsened, accompanied by a significant increase in serum transaminases, inflammatory responses, oxidative stress, and TUNEL staining. Strikingly, ECMO not only stabilized hemodynamics after DBD but also mitigated liver damage, leading to an alleviated status post liver transplantation. ECMO stabilizes hemodynamics, attenuates inflammatory responses and oxidative stress, thereby enhancing the quality of liver grafts for transplantation.

Evaluation of Minimum-to-Severe Global and Macrovesicular Steatosis in Human Liver Specimens: A Portable Ambient Light-Compatible Spectroscopic Probe.

Hepatic steatosis (HS), particularly macrovesicular steatosis (MaS), influences transplant outcomes. Accurate assessment of MaS is crucial for graft selection. While traditional assessment methods have limitations, non-invasive spectroscopic techniques like Raman and reflectance spectroscopy offer promise. This study aimed to evaluate the efficacy of a portable ambient light-compatible spectroscopic system in assessing global HS and MaS in human liver specimens. A two-stage approach was employed on thawed snap-frozen human liver specimens under ambient room light: biochemical validation involving a comparison of fat content from Raman and reflectance intensities with triglyceride (TG) quantifications and histopathological validation, contrasting Raman-derived fat content with evaluations by an expert pathologist and a "Positive Pixel Count" algorithm. Raman and reflectance intensities were combined to discern significant (≥ 10%) discrepancies in global HS and MaS. The initial set of 16 specimens showed a positive correlation between Raman and reflectance-derived fat content and TG quantifications. The Raman system effectively differentiated minimum-to-severe global and macrovesicular steatosis in the subsequent 66 specimens. A dual-variable prediction algorithm was developed, effectively classifying significant discrepancies (> 10%) between algorithm-estimated global HS and pathologist-estimated MaS. Our study established the viability and reliability of a portable spectroscopic system for non-invasive HS and MaS assessment in human liver specimens. The compatibility with ambient light conditions and the ability to address limitations of previous methods marks a significant advancement in this field. By offering promising differentiation between global HS and MaS, our system introduces an innovative approach to real-time and quantitative donor HS assessments. The proposed method holds the promise of refining donor liver assessment during liver recovery and ultimately enhancingtransplantation outcomes.

Impact of donor organ quality on recipient outcomes in lung transplantation: 14-Year single-center experience using the Eurotransplant lung donor score

Impact of donor organ quality on recipient outcomes in lung transplantation: 14-Year single-center experience using the Eurotransplant lung donor score

Impact of Clinical and Pharmacological Parameters on Faecal Microbiota Transplantation Outcome in Clostridioides difficile Infections: Results of a 5-Year French National Survey.

Detailed comparative assessment of procedure-related factors associated with faecal microbiota transplantation (FMT) efficacy in Clostridioides difficile infection (CDI) is limited. We took advantage of the differences in procedures at the various French FMT centres to determine clinical and procedure-related factors associated with FMT success in CDI. We performed a nationwide retrospective multicentre cohort study. All FMTs performed within The French Faecal Transplant Group for CDI from 2018 to 2022 were included. Clinical data were collected retrospectively from recipient medical files, characteristics of stool transplant preparations were prospectively collected by each Pharmacy involved. Univariate and multivariate analyses were performed using Fisher's test and multiple logistic regression. Six hundred fifty-eight FMTs were performed for 617 patients in 17 centres. The overall efficacy of FMT was 84.3% (520/617), with 0.5% of severe adverse events possibly related to FMT (3/658). Forty-seven patients were treated at the first recurrence of CDI with a similar success rate (85.1%). Severe chronic kidney disease (CKD; OR: 2.18, 95%CI [1.20-3.88]), non-severe refractory CDI (OR: 15.35, [1.94-318.2]), the use of ≥ 80% glycerol (OR: 2.52, [1.11-5.67]), insufficient bowel cleansing (OR: 5.47, [1.57-20.03]) and partial FMT retention (OR: 9.97, [2.62-48.49]) were associated with CDI recurrence within 8 weeks. Conditions of transplant manufacturing, bowel cleansing, and a route of delivery tailored to the patient's characteristics are key factors in optimising FMT efficacy. FMT at first recurrence showed high success in real-life practice, whereas it had lower efficacy in severe CDI and non-severe refractory CDI.

Effects of Root Pruning and Size on Growth Traits of Hybrid Poplar Seedlings

Selecting seedlings of varying sizes and effectively managing root pruning are key challenges in transplantation. However, the effects of seedling size and root pruning on transplantation outcomes are not fully understood. This study classified one-year-old Populus ‘Beilinxiongzhu-01’ seedlings into three size categories based on height: large (308.75 ± 9.66 cm), medium (238.00 ± 7.71 cm), and small (138.92 ± 7.18 cm). In early March of the subsequent year, root pruning was applied with varying intensities based on root collar diameter: low (15 times), medium (7.5 times), and high (3.75 times). A control group without pruning was also included. Over the year, key phenological and morphological traits were monitored. The results showed that (1) root pruning significantly impacted the phenology of seedlings, accelerating root emergence, delaying early leaf phenology, increasing the dieback rate, and postponing end-of-season defoliation. Mortality and the rapid growth phase were not significantly affected. Larger seedlings exhibited earlier end-of-season defoliation and higher dieback rates early in the growing season, while smaller seedlings advanced in early leaf development. (2) Except under low or no pruning, root pruning reduced seedling height (H), diameter at breast height (DBH), and root collar diameter (RCD). However, across all treatments, these indicators remained higher in larger seedlings compared to smaller ones. Under medium- and high-intensity pruning, smaller seedlings exhibited higher relative growth rates and larger leaf areas than larger seedlings, with the reduction in these variables becoming more pronounced as seedlings increased in size. Notably, only larger seedlings demonstrated a reduction in maximum growth rate, suggesting greater vulnerability to root pruning. In summary, root pruning induced significant phenological and morphological differences across seedling sizes. While smaller seedlings showed some response to pruning, larger seedlings experienced more pronounced phenological disruptions and growth inhibition.