Ex vivo culture resting time impacts transplantation outcomes of genome-edited human hematopoietic stem and progenitor cells in xenograft mouse models.

Ex vivo resting culture is a standard procedure following genome editing in hematopoietic stem and progenitor cells (HSPCs). However, prolonged culture may critically affect cell viability and stem cell function. We investigated whether varying durations of culture resting times impact the engraftment efficiency of human CD34+ HSPCs edited at the BCL11A enhancer, a key regulator in the expression of fetal hemoglobin. We employed electroporation to introduce CRISPR-Cas9 components for BCL11A enhancer editing and compared outcomes with nonelectroporated (NEP) and electroporated-only (EP) control groups. Post-electroporation, we monitored cell viability, death rates, and the frequency of enriched hematopoietic stem cell (HSC) fractions (CD34+CD90+CD45RA- cells) over a 48-hour period. Our findings reveal that while the NEP group showed an increase in cell numbers 24 hours post-electroporation, both EP and BCL11A-edited groups experienced significant cell loss. Although CD34+ cell frequency remained high in all groups for up to 48 hours post-electroporation, the frequency of the HSC-enriched fraction was significantly lower in the EP and edited groups compared to the NEP group. In NBSGW xenograft mouse models, both conditioned with busulfan and nonconditioned, we found that immediate transplantation post-electroporation led to enhanced engraftment without compromising editing efficiency. Human glycophorin A+ (GPA+) red blood cells (RBCs) sorted from bone marrow of all BCL11A edited mice exhibited similar levels of γ-globin expression, regardless of infusion time. Our findings underscore the critical importance of optimizing the culture duration between genome editing and transplantation. Minimizing this interval may significantly enhance engraftment success and minimize cell loss without compromising editing efficiency. These insights offer a pathway to improve the success rates of genome editing in HSPCs, particularly for conditions like sickle cell disease.

cMPL-Based Purification and Depletion of Human Hematopoietic Stem Cells: Implications for Pre-Transplant Conditioning.

The transplantation of gene-modified autologous hematopoietic stem and progenitor cells (HSPCs) offers a promising therapeutic approach for hematological and immunological disorders. However, this strategy is often limited by the toxicities associated with traditional conditioning regimens. Antibody-based conditioning strategies targeting cKIT and CD45 antigens have shown potential in mitigating these toxicities, but their long-term safety and efficacy in clinical settings require further validation. In this study, we investigate the thrombopoietin (TPO) receptor, cMPL, as a novel target for conditioning protocols. We demonstrate that high surface expression of cMPL is a hallmark feature of long-term repopulating hematopoietic stem cells (LT-HSCs) within the adult human CD34+ HSPC subset. Targeting the cMPL receptor facilitates the separation of human LT-HSCs from mature progenitors, a delineation not achievable with cKIT. Leveraging this finding, we developed a cMPL-targeting immunotoxin, demonstrating its ability to selectively deplete host cMPLhigh LT-HSCs with a favorable safety profile and rapid clearance within 24 hours post-infusion in rhesus macaques. These findings present significant potential to advance our understanding of human hematopoiesis and enhance the therapeutic outcomes of ex vivo autologous HSPC gene therapies.

Influence of the menstrual cycle on static and dynamic kinematics of the foot medial longitudinal arch.

BACKGROUND: In women, the laxity of the plantar fascia increases during the ovulation phase of the menstrual cycle. Although it is possible that this increased laxity results in a decreased height of the foot in the medial longitudinal arch and exacerbates symptoms of several overuse injuries of the lower extremity, the influence of the menstrual cycle on static and dynamic kinematics of the medial longitudinal arch is unclear. The purpose of this study was to confirm that the medial longitudinal arch height during static standing, gait, and landing decrease during the menstrual cycle ovulation phase. METHODS: Participants in this study were 16 female college students with normal menstrual cycles and 16 male college students. Navicular height in the static standing position was measured using a three-dimensional foot scanner. Kinematics of the medial longitudinal arch during gait and landing were measured using a three-dimensional motion capture system to determine the navicular height at initial contact, minimal navicular height, and dynamic navicular drop. In all measurements, female participants were tested twice during the course of one complete menstrual cycle: once during the follicular phase and once during the ovulation phase. Male participants were tested twice with an interval of ≥1 week and <2 weeks. RESULTS: In women, navicular height in the static standing position significantly decreased during the ovulation phase compared with follicular phase (mean difference [95% confidence interval] = 2.1 [0.9-3.4] mm; p = 0.002), whereas men showed no statistical difference between the first and second measurements. In both men and women, no statistical differences were identified for the dynamic medial longitudinal arch kinematics measured during gait and landing. CONCLUSIONS: Navicular height in the static standing position slightly decreased during the ovulation phase.

Post-Transplant Administration of G-CSF Impedes Engraftment of Gene Edited Human Hematopoietic Stem Cells by Exacerbating the p53-Mediated DNA Damage Response.

Granulocyte colony stimulating factor (G-CSF) is commonly used as adjunct treatment to hasten recovery from neutropenia following chemotherapy and autologous transplantation of hematopoietic stem and progenitor cells (HSPCs) for malignant disorders. However, the utility of G-CSF administration after ex vivo gene therapy procedures targeting human HSPCs has not been thoroughly evaluated. Here, we provide evidence that post-transplant administration of G-CSF impedes engraftment of CRISPR-Cas9 gene edited human HSPCs in xenograft models. G-CSF acts by exacerbating the p53-mediated DNA damage response triggered by Cas9- mediated DNA double-stranded breaks. Transient p53 inhibition in culture attenuates the negative impact of G-CSF on gene edited HSPC function. In contrast, post-transplant administration of G-CSF does not impair the repopulating properties of unmanipulated human HSPCs or HSPCs genetically engineered by transduction with lentiviral vectors. The potential for post-transplant G-CSF administration to aggravate HSPC toxicity associated with CRISPR-Cas9 gene editing should be considered in the design of ex vivo autologous HSPC gene editing clinical trials.

Modulation of WNT, Activin/Nodal, and MAPK Signaling Pathways Increases Arterial Hemogenic Endothelium and Hematopoietic Stem/Progenitor Cell Formation During Human iPSC Differentiation.

Several differentiation protocols enable the emergence of hematopoietic stem and progenitor cells (HSPCs) from human-induced pluripotent stem cells (iPSCs), yet optimized schemes to promote the development of HSPCs with self-renewal, multilineage differentiation, and engraftment potential are lacking. To improve human iPSC differentiation methods, we modulated WNT, Activin/Nodal, and MAPK signaling pathways by stage-specific addition of small-molecule regulators CHIR99021, SB431542, and LY294002, respectively, and measured the impact on hematoendothelial formation in culture. Manipulation of these pathways provided a synergy sufficient to enhance formation of arterial hemogenic endothelium (HE) relative to control culture conditions. Importantly, this approach significantly increased production of human HSPCs with self-renewal and multilineage differentiation properties, as well as phenotypic and molecular evidence of progressive maturation in culture. Together, these findings provide a stepwise improvement in human iPSC differentiation protocols and offer a framework for manipulating intrinsic cellular cues to enable de novo generation of human HSPCs with functionality in vivo.

Fibrin clot and Leukocyte-rich platelet-rich fibrin show similar release kinetics and amount of growth factors: a pilot study.

BACKGROUND: In knee arthroscopic surgery, fibrin clot (FC) and leukocyte-rich platelet-rich fibrin (L-PRF) may be used in augmentation for meniscal repair. Studies have investigated growth factors released from FC and L-PRF; however, it is difficult to compare FC and L-PRF between different studies. Direct comparison of growth factors that may support meniscal healing released from FC and L-PRF may be beneficial in deciding whether to use FC or L-PRF. If no significant difference is seen, the surgeon may decide to use FC which is easier to prepare compared to L-PRF. The purpose of this pilot study is to investigate the release amount and pattern of basic fibroblast growth factor (bFGF), platelet-derived growth factor AB (PDGF-AB), transforming growth factor ß1 (TGF-ß1), vascular endothelial growth factor (VEGF), and stromal cell-derived factor 1 (SDF-1) from FC and L-PRF. METHOD: Twenty milliliters (ml) of whole blood was collected from each of the four volunteers. Ten milliliters of whole blood was allocated for preparation of FC and 10 ml for L-PRF. FC and L-PRF were separately placed in 5 ml of culture media. Five milliliters of the culture media was sampled and refilled at 15 min, 1 day, 3 days, 1 week and 2 weeks. The collected culture was used to quantify bFGF, PDGF-AB, TGF-ß1, VEGF, and SDF-1 release by Enzyme-linked immune-sorbent assay (ELISA). Mann-Whitney U test was performed to assess significance of differences in amount of each growth factor released between FC and L-PRF. Significance was accepted at P value less than 0.05. RESULTS: At two weeks, the cumulative release of TGF-ß1 was the highest among all the growth factors in both FC and L-PRF (FC:19,738.21 pg/ml, L-PRF: 16,229.79 pg/ml). PDGF-AB (FC: 2328 pg/ml, L-PRF 1513.57 pg/ml) had the second largest amount, followed by VEGF (FC: 702.06 pg/ml, L-PRF 595.99 pg/ml) and bFGF (FC: 23.48 pg/ml, L-PRF 18.2 pg/ml), which order was also common in both FC and L-PRF. No significant difference in final release amount and pattern was seen between FC and L-PRF. CONCLUSION: The current pilot study showed that cumulative release amount and release pattern of PDGF-AB, VEGF, TGF-ß1, and bFGF did not significantly differ between FC and L-PRF during the two weeks of observation.

Modulation of WNT, Activin/Nodal and MAPK Signaling Pathways Increases Arterial Hemogenic Endothelium and Hematopoietic Stem/Progenitor Cell Formation During Human iPSC Differentiation.

Several differentiation protocols enable the emergence of hematopoietic stem and progenitor cells (HSPCs) from human induced pluripotent stem cells (iPSCs), yet optimized schemes to promote the development of HSPCs with self-renewal, multilineage differentiation and engraftment potential are lacking. To improve human iPSC differentiation methods, we modulated WNT, Activin/Nodal and MAPK signaling pathways by stage-specific addition of small molecule regulators CHIR99021, SB431542 and LY294002, respectively, and measured the impact on hematoendothelial formation in culture. Manipulation of these pathways provided a synergy sufficient to enhance formation of arterial hemogenic endothelium (HE) relative to control culture conditions. Importantly, this approach significantly increased production of human HSPCs with self-renewal and multilineage differentiation properties, as well as phenotypic and molecular evidence of progressive maturation in culture. Together, these findings provide a stepwise improvement in human iPSC differentiation protocols and offer a framework for manipulating intrinsic cellular cues to enable de novo generation of human HSPCs with functionality in vivo . Significance Statement: The ability to produce functional HSPCs by differentiation of human iPSCs ex vivo holds enormous potential for cellular therapy of human blood disorders. However, obstacles still thwart translation of this approach to the clinic. In keeping with the prevailing arterial-specification model, we demonstrate that concurrent modulation of WNT, Activin/Nodal and MAPK signaling pathways by stage-specific addition of small molecules during human iPSC differentiation provides a synergy sufficient to promote arterialization of HE and production of HSPCs with features of definitive hematopoiesis. This simple differentiation scheme provides a unique tool for disease modeling, in vitro drug screening and eventual cell therapies.

Talar component subsidence correlates with periprosthetic osteolysis after total ankle arthroplasty.

PURPOSE: This study aimed to investigate the relationship between periprosthetic osteolysis around the talar component and the amount of talar component subsidence after total ankle arthroplasty (TAA). METHODS: This study included forty patients who underwent TAA with a mean follow-up of 67.5 ± 17.0 months. The patients were divided into two groups based on the amount of osteolysis around the talar component, as measured by computed tomography at the latest clinic visit: none to 2 mm (N group, n = 20) and greater than or equal to 2 mm (O group, n = 20). The average amount of talar component subsidence, clinical outcomes, and complications were compared between the two groups. In the O group, the correlation between osteolysis and talar component subsidence was evaluated. RESULTS: The average talar component subsidence was significantly different between the N (0.22 ± 0.94 mm) and O groups (2.12 ± 2.28 mm). Five out of 20 ankles in the O group required revision surgery owing to talar component subsidence. The Japanese Society for Surgery of the Foot scores in the N and O groups were significantly different: 93.5 ± 7.7 and 85.3 ± 15.4, respectively. In the O group, we found that osteolysis tended to develop on the lateral side, and the amount of osteolysis was positively correlated with the talar component subsidence (r = 0.59, P = .007). CONCLUSION: In the O group, a positive correlation between osteolysis and talar component subsidence was found, and five patients required revision surgery.

Risk factors of residual pivot-shift after anatomic double-bundle anterior cruciate ligament reconstruction.

INTRODUCTION: Although anterior cruciate ligament reconstruction (ACLR) is considered a successful procedure, residual pivot-shift after surgery remains to be solved. The purpose of this study was to comprehensively evaluate the risk factors of residual pivot-shift after anatomic double-bundle (DB) ACLR. MATERIALS AND METHODS: A total of 164 patients who underwent primary anatomic DB-ACLR between January 2014 and December 2019 and screw removal after the index ACLR in our hospital were included in this retrospective case-control study. The manual pivot-shift test was performed under general anesthesia during screw removal surgery, and patients with grade 1 or higher pivot-shift were classified as the positive pivot-shift group, and those with grade 0 were defined as the negative pivot-shift group. Univariate and logistic regression analyses were performed to identify the factors associated with postoperative residual pivot-shift. Assessment included sex, age, time to surgery, preoperative Tegner activity scale, preoperative pivot-shift grade, preoperative anterior tibial translation by the KT-2000 arthrometer measurement, meniscus injury and its surgical procedure, knee hyperextension, cartilage damage, Segond fracture, medial and lateral posterior tibial slope, lateral-medial slope asymmetry, participation in pivoting sport/activity at the time of injury, and return to sports at postoperative one year line. RESULTS: Postoperative positive pivot-shift was observed in 14 (8.5%) of 164 patients. The KT-2000 measurement at 1-year postoperatively was significantly higher in the residual pivot-shift-positive group than in the negative group (P < 0.05). Logistic regression analysis revealed that age of patients < 20 years [P < 0.05, odds ratio (OR): 6.1)], preoperative pivot-shift grade (P < 0.05, OR: 4.4), and hyperextended knee (P < 0.05, OR: 11.8) were risk factors of postoperative pivot-shift. There were no statistically significant differences between other variables. CONCLUSIONS: Patients < 20 years of age, with high-grade preoperative pivot-shift, or hyperextended knees had a higher risk of residual postoperative pivot-shift.

Short-Term Clinical Outcomes Following Total Ankle Arthroplasty Without Concomitant Osteotomy in Ankles with Severe Preoperative Varus Deformity: Comparison to Ankles with Preoperative Neutral Alignment.

BACKGROUND: It remains controversial whether satisfactory outcomes can be obtained following total ankle arthroplasty (TAA) without osteotomy in patients with severe varus ankle deformities. This study aimed to examine outcomes following TAA without concomitant osteotomies in patients with severe varus ankle alignment by comparing them with those in patients with neutral alignment. METHODS: Fifty-one patients (53 ankles; mean age, 71.4 ± 5.6 years) who underwent TAA using the TNK ankle prosthesis were examined (mean follow-up, 36.8 ± 17.8 months). Patients were allocated into groups according to the preoperative talar tilt (TT) angle: the neutral group (preoperative TT angle <10°; n = 37) and the varus group (preoperative TT angle ≥10°; n = 16). Outcome measures, including the Japanese Society for Surgery of the Foot scale, Self-Administered Foot Evaluation Questionnaire, ankle range of motion, and radiographic parameters, were assessed before surgery and at the final follow-up. RESULTS: Significant improvements were observed in clinical and radiographic outcomes in both groups after surgery. Postoperative Japanese Society for Surgery of the Foot scale and subscale scores of pain and shoes in the Self-Administered Foot Evaluation Questionnaire were not significantly different between the groups, whereas subscale scores of function, social, and health were greater in the varus group than in the neutral group at the final follow-up. Radiographic parameters, including TT angle and tibial axis-medial malleolus (TMM) angle, improved postoperatively and were not significantly different between the neutral (mean TT angle, 0.5 ± 0.7°; mean TMM angle, 16.0 ± 4.6°) and varus (meanTT angle, 0.4 ± 0.7°; meanTMM angle, 17.0 ± 5.3°) groups at the final follow-up. To achieve neutral alignment, adjunctive procedures were required more often in the varus group. CONCLUSIONS: Outcomes of TAA using the TNK ankle prosthesis were favorable in patients with severe varus ankle and in those with neutral ankle without concomitant osteotomy. Satisfactory outcomes could be achieved in patients with severe varus ankle alignment after TAA without concomitant osteotomy.

Influence of Injury to the Kaplan Fibers of the Iliotibial Band on Anterolateral Rotatory Knee Laxity in Anterior Cruciate Ligament Injury: A Retrospective Cohort Study.

BACKGROUND: Biomechanical cadaveric studies have shown that Kaplan fibers (KFs) of the iliotibial band play a role in controlling anterolateral rotatory knee laxity in anterior cruciate ligament (ACL) injury. However, in the clinical setting, the contribution of injury to KFs on anterolateral rotatory laxity remains unclear. PURPOSE: To use magnetic resonance imaging (MRI) scans to detect concomitant KF injury in ACL-injured knees and to then examine the effect of KF injury on anterolateral rotatory laxity as measured by the pivot-shift test in a clinical setting. STUDY DESIGN: Cross-sectional study; Level of evidence, 3. METHODS: The study enrolled 91 patients with primary ACL tears (mean age 25 ± 11 years; 46 male and 45 female) whose MRI was conducted within 90 days after injury. KF injury was assessed by MRI according to previously reported criteria, and the patients were allocated to a KF injury group and a no-KF injury group. At the time of ACL reconstruction, the pivot-shift test was performed with the patient under anesthesia and quantitatively evaluated by tibial acceleration using an electromagnetic measurement system. Manual grading of the pivot-shift test was assessed according to guidelines of the International Knee Documentation Committee. The data were statistically compared between the 2 groups using Mann-Whitney U test and Fisher exact test (P < .05). RESULTS: KFs were identified in 85 patients (93.4%), and KF injury was detected in 20 of the 85 patients (23.5%). No significant differences were observed between the KF injury group (n = 20) and the no-KF injury group (n = 65) in demographic characteristics, the period from injury to MRI (8.0 ± 14.0 days vs 8.9 ± 12.1 days, respectively), the rate of meniscal injury (50.0% vs 53.8%), or the rate of anterolateral ligament injury (45.0% vs 44.6%). Regarding the pivot-shift test, no significant differences were observed in tibial acceleration (1.2 m/s2 [interquartile range, 0.5-2.1 m/s2] vs 1.0 m/s2 [interquartile range, 0.6-1.7 m/s2], respectively) or manual grading between the 2 groups. CONCLUSION: Concomitant KF injury did not significantly affect the pivot-shift phenomenon in acute ACL-injured knees. The findings suggest that the contribution of KF injury to anterolateral rotatory knee laxity may be limited in the clinical setting.

Analysis of popliteal artery location for high tibial and distal tuberosity osteotomy using contrast-enhanced computed tomography.

BACKGROUND: Our objective was to evaluate the location of popliteal artery (PA) in osteotomy planes during high tibial osteotomy (HTO) and to determine a safer angle for screw drilling to the tibial tuberosity during distal tuberosity osteotomy (DTO). METHODS: Twenty knees in 20 patients who underwent contrast-enhanced computed tomography for cardiovascular diseases were examined. Osteotomy planes for open-wedge HTO (OWHTO) and hybrid closed-wedge HTO (hybrid CWHTO) were created using three-dimensional bone models. The distance from the posterior cortex of the tibia to the PA (dPC-PA) in the osteotomy planes was measured in the virtual osteotomy planes. The dangerous point (Point D1) was defined as the point 17.5 mm away from PA, setting the working length of the bone saw as 35 mm. The distance between the most medial point of the tibial cortex (Point M) and Point D1 in OWHTO and the most lateral point (Point L) and Point D1 in hybrid CWHTO were examined (dM-D1 and dL-D1, respectively). The location of Point D1 to the osteotomy line (%D1) was expressed as percentage, setting the start and end of the osteotomy line as 0% and 100%, respectively. To determine the safe angle for screw drilling in DTO, the angle between the line tangential to the medial cortex of the tibia and that passing through the center of the tibial tuberosity and PA were measured. RESULTS: In OWHTO and hybrid CWHTO, the mean dPC-PA was 10.6 mm (6.9-16.5 mm) and 10.2 mm (7.3-15.4 mm), respectively. The mean dM-D1 in OWHTO was 25.9 mm (24.6-27.2 mm) and dL-D1 in hybrid CWHTO was 5.1 mm (2.9-7.4 mm). The mean %D1 was 47.6 ± 3.7% in OWHTO and 9.3 ± 4.1% in hybrid CWHTO, respectively. The minimal angle between the two lines in DTO was 35.2°. CONCLUSION: PAs could run within 10 mm from the posterior cortex in the osteotomy planes of HTO. Therefore, proper posterior protection is necessary when cutting posterior cortex. An angle of less than 35° against the medial cortex line would be safe for screw fixation to avoid vascular injury in DTO.

A phase I/IIa clinical trial of third-generation autologous chondrocyte implantation (IK-01) for focal cartilage injury of the knee.

Background/objective: The purpose of this study was to report the outcomes of a clinical trial conducted in Japan to assess the safety and effectiveness of third-generation autologous chondrocyte implantation (ACI) using IK-01 (CaReS™), which does not require flap coverage, in the treatment of patients with focal cartilage injury of the knee. Methods: This was an open label, exploratory clinical trial. Patients were enrolled between June 2012 and September 2016. The primary endpoint of the study was the International Knee Documentation Committee (IKDC) score at 52 weeks after implantation. The IKDC, Lysholm, and visual analog scale (VAS) scores were evaluated at the time of screening and at 4, 12, 24, 36, and 52 weeks after implantation. Improvements from the baseline scores were evaluated using the equation "(postoperative score) - (preoperative score)." Magnetic resonance imaging (MRI) was performed at 2, 12, 24, and 52 weeks after implantation, and MRI measurements were evaluated using T1 rho and T2 mapping. Results: Nine patients were enrolled in this study and were examined for safety. Product quality did not satisfy the specification in one patient, and bacterial joint infection occurred in one patient. As a result, seven patients were included in the outcome analyses. The mean IKDC score significantly improved from 36.4 preoperatively to 74.1% at 52 weeks after implantation (p < 0.0001). The mean Lysholm and VAS scores also significantly improved from 39.6 to 57.4 to 89.6 and 22.9, respectively, after surgery. In the MRI evaluation, the T1 rho and T2 values of the implanted area were similar to those of the surrounding cartilage at 52 weeks after implantation. Conclusions: Third generation ACI (IK-01) can be an effective treatment option for focal cartilage defects of the knee; however, surgeons must pay careful attention to the risk of postoperative joint infection.

Direct suturing quadriceps tendon to a continuous loop with a suspensory button provides biomechanically superior fixation in ACL reconstruction.

PURPOSE: To compare the biomechanical strength of different fixation configurations using suspensory buttons in a soft-tissue quadriceps tendon (QT) grafts in anterior cruciate ligament (ACL) reconstruction. METHODS: Forty bovine QTs, 6-cm long and 10-mm wide, were allocated into four groups with different suture configurations using suspensory buttons (n = 10 in each group): Group A, a baseball suture with a knot tied to the continuous loop with a suspensory button; Group B, same configuration as in Group A but with the knot tied at the opposite end of the baseball suture; Group C, a continuous loop with a suspensory button stitched directly to the QT with simple sutures, and Group D, a baseball suture tied directly to a suspensory button. Biomechanical testing was performed by preloading followed by cyclic loading for 500 cycles between 10 and 100 N. The length of elongation (mm) and maximum load to failure (N) were recorded, and compared among the four groups. RESULTS: Group C showed significantly smaller elongation (4.1 mm [95% CI 3.1-5.2]) than Group A (8.2 mm [95% CI 7.0-9.4]), Group B (10.5 mm [95% CI 7.7-13.3]), and Group D (8.5 mm [95% CI 7.0-9.9]) (A-C; P = 0.004, B-C; P = 0.0001, C-D; P = 0.0018). The maximum load to failure in Group C (386 N [95%CI 306-466]) was significantly higher than that in Group A (196 N [95% CI 141-251]), Group B (226 N [95% CI 164-289]), and Group D (212 N [95%CI 171-253]) (A-C; P = 0.0001, B-C; P = 0.0009, C-D; P = 0.0002). No significant differences were observed between Group A, B, and D in terms of elongation and maximum load to failure. CONCLUSION: The soft-tissue QT graft fixation configuration stitched directly to a continuous loop with suspensory button using simple sutures exhibits small elongation and high maximum load to failure among the four configurations. Regarding clinical relevance, direct suturing of the soft-tissue QT to a continuous loop with a suspensory button may be advantageous for femoral fixation in ACL reconstruction from a biomechanical perspective, and warrant future development of a novel fixation device using this principle.

Satisfaction with playing pre-injury sports 1 year after anterior cruciate ligament reconstruction using a hamstring autograft.

BACKGROUND: Few studies have examined patient satisfaction with playing pre-injury sports after anterior cruciate ligament (ACL) reconstruction. The purpose of this study was to investigate patient satisfaction with playing pre-injury sport and identify factors associated with satisfaction. METHODS: A total of 97 patients underwent unilateral ACL reconstruction using a hamstring autograft and returned to pre-injury sports 1 year after surgery. Patient satisfaction with playing pre-injury sport was assessed by a visual analog scale (VAS) and an ordinal four-grade scale. Problems related to the operated knee were also assessed. Knee muscle strength, single leg hop distance, knee laxity, subjective knee pain, and fear of movement/reinjury using Tampa Scale for Kinesiophobia-11 (TSK-11) were measured. Multivariate linear regression analysis was performed to determine the factors associated with patient satisfaction with playing pre-injury sport 1 year after surgery. RESULTS: The average VAS score for patient satisfaction with playing pre-injury sports 1 year after surgery was 77.8 ± 20.2. Of the 97 patients, 87 patients (89.7%) answered "satisfied" or "mostly satisfied", whereas 51 patients (52.6%) had one or more problems. Multivariate linear regression analysis identified that the TSK-11 score was associated with patient satisfaction with playing a pre-injury sport 1 year after surgery. CONCLUSION: Most of the patients who returned to pre-injury sports were satisfied with their outcomes. In contrast, approximately half of the patients had one or more problems after returning to play pre-injury sports. In particular, fear of movement/reinjury was significantly associated with patient satisfaction with playing pre-injury sport 1 year after surgery.

NOTCH-mediated ex vivo expansion of human hematopoietic stem and progenitor cells by culture under hypoxia.

Activation of NOTCH signaling in human hematopoietic stem/progenitor cells (HSPCs) by treatment with an engineered Delta-like ligand (DELTA1ext-IgG [DXI]) has enabled ex vivo expansion of short-term HSPCs, but the effect on long-term repopulating hematopoietic stem cells (LTR-HSCs) remains uncertain. Here, we demonstrate that ex vivo culture of human adult HSPCs with DXI under low oxygen tension limits ER stress in LTR-HSCs and lineage-committed progenitors compared with normoxic cultures. A distinct HSC gene signature was upregulated in cells cultured with DXI in hypoxia and, after 21 days of culture, the frequency of LTR-HSCs increased 4.9-fold relative to uncultured cells and 4.2-fold compared with the normoxia + DXI group. NOTCH and hypoxia pathways intersected to maintain undifferentiated phenotypes in cultured HSPCs. Our work underscores the importance of mitigating ER stress perturbations to preserve functional LTR-HSCs in extended cultures and offers a clinically feasible platform for the expansion of human HSPCs.

Effect of Percentage of Femoral Anterior Cruciate Ligament Insertion Site Reconstructed With Hamstring Tendon on Knee Kinematics and Graft Force.

BACKGROUND: Previous studies have stated that closely matching the size of the anterior cruciate ligament (ACL) insertion site footprint is important for biomechanical function and clinical stability after ACL reconstruction. However, the ACL varies widely regarding the area of femoral insertion, tibial insertion, and midsubstance of ACL, and reconstructing the insertion site area with a uniform diameter graft can result in a cross-sectional area that is greater than that of the midsubstance of the native ACL. Therefore, understanding the effect of relative graft size in ACL reconstruction on knee biomechanics is important for surgical planning. PURPOSE: To assess how the percentage of femoral insertion site affects knee biomechanics in single- and double-bundle ACL reconstruction. STUDY DESIGN: Controlled laboratory study. METHODS: A total of 14 human cadaveric knees were scanned with magnetic resonance imaging and tested using a robotic system under an anterior tibial load and a combined rotational load. In total, 7 knee states were evaluated: intact ACL; deficient ACL; single-bundle ACL reconstruction with approximate graft sizes 25% (small), 50% (medium), and 75% (large) of the femoral insertion site; and double-bundle reconstruction of approximately 50% (medium) and 75% (large) of the femoral insertion site, based on the ratio of the cross-sectional area of the graft to the area of the femoral ACL insertion site determined by magnetic resonance imaging. RESULTS: Anterior tibial translation was not significantly larger than the intact state in single-bundle and double-bundle medium graft reconstructions (P > .05) and was significantly greater in the single-bundle small graft reconstruction (P < .05). Anterior knee translation in single-bundle medium graft and large graft reconstructions was not statistically different (P > .05). In contrast, the anterior tibial translation for double-bundle large graft reconstruction was significantly smaller than for double-bundle medium graft reconstruction at low flexion angles (P < .05). The single-bundle small graft force was significantly different from the intact ACL in situ force (P < .05). The graft force with double-bundle large reconstruction was significantly greater than that with the double-bundle medium reconstruction (P < .05) but was not significantly different from that of the intact ACL (P > .05). CONCLUSION: Knee biomechanics with a single-bundle small graft tended to be significantly different from those of the intact knee. In the kinematic and kinetic data for the single- and double-bundle medium graft reconstruction, only the anterior translation at full extension for the single-bundle reconstruction was significantly different (lower) from that of intact knee. This was a time zero study. CLINICAL RELEVANCE: This study can provide surgeons with guidance in selecting the graft size for ACL reconstruction.

Differences in preoperative planning for high-tibial osteotomy between the standing and supine positions.

INTRODUCTION: Previous studies have reported that alignment changes depend on the patient's position in orthopedic surgery. However, it has not yet been well examined how the patient's position affects the preoperative planning in high-tibial osteotomy (HTO). Therefore, the aim of this study was to investigate the effects of the patient's position on preoperative planning in HTO. MATERIALS AND METHODS: A total of 60 knees in 55 patients who underwent HTO were retrospectively examined. Virtual preoperative planning for medial open-wedge HTO (OWHTO), lateral closed-wedge HTO (CWHTO), and hybrid CWHTO were performed by setting the percentage of the weight-bearing line (%WBL) at 62% as an optimal alignment. The correction angle differences between the supine and standing radiographs were measured. The virtual %WBL (v%WBL) was determined by applying the correction angle obtained from the standing radiograph to the supine radiograph. The %WBL discrepancy (%WBLd) was calculated as v%WBL - 62 (%) to predict the possible correction errors during surgeries. A single regression analysis was performed to examine the correlation between the correction angle difference and %WBLd. RESULTS: The mean correction angle was significantly higher when the preoperative planning was based on standing radiographs than when based on supine radiographs (P < 0.001), and the mean difference was 2.2 ± 1.5°. The difference between the two conditions in the medial opening gaps for OWHTO, lateral wedge sizes (mm) for CWHTO, and hybrid CWHTO were 2.6 ± 2.0, 2.3 ± 1.6, and 1.9 ± 1.4, respectively. The mean v%WBL was 71.2% ± 7.3%, and the mean %WBLd was 10.1% ± 7.4%. A single regression analysis revealed a linear correlation between the correction angle difference and %WBLd (%WBLd = 4.72 × correction angle difference + 0.08). No statistically significant difference in the parameters was found between the supine and standing radiographs postoperatively. CONCLUSIONS: We found significant differences in the estimated correction angles between the supine and standing radiographs in the planning for HTO. Therefore, surgeons should carefully consider the difference between supine and standing radiographs and estimate the possible correction error during surgery when planning a HTO.

Influence of selected plane on the evaluation of tibial tunnel locations using a three-dimensional bone model in double-bundle anterior cruciate ligament reconstruction.

BACKGROUND: The purpose of this study was to investigate the influence of a selected plane on the evaluation of tibial tunnel locations following anterior cruciate ligament reconstruction (ACLR) between two planes: the plane parallel to the tibial plateau (Plane A) and the plane perpendicular to the proximal tibial shaft axis (Plane B). METHODS: Thirty-four patients who underwent double-bundle ACLR were included. Three-dimensional model of tibia was created using computed tomography images 2 weeks postoperatively, and tibial tunnels of the anteromedial bundle (AMB) and posterolateral bundle (PLB) were extracted. To evaluate tibial tunnel locations, two planes (Planes A and B) were created. The locations of the tibial tunnel apertures of each bundle were evaluated using a grid method and compared between Planes A and B. The difference in coronal alignment between Planes A and B were also assessed. RESULTS: The AMB and PLB tunnel apertures in Plane A were significantly more laterally located than in Plane B (mean difference; AMB, 1.5%; PLB, 1.7%, P < 0.01). There were no significant differences in the anteroposterior direction between the planes. Coronal alignment difference between the planes was 16.8 ± 2.2°; Plane B was more valgus than Plane A. CONCLUSION: Although tibial tunnel locations were not significantly influenced by the selected planes in the AP direction, subtle but statistically significant differences were found in the ML direction between the Planes A and B in double-bundle anterior cruciate ligament reconstruction. The findings suggest that both Planes A and B can be used in the assessment of tibial tunnel locations after anterior cruciate ligament reconstruction.

Anatomic double-bundle anterior cruciate ligament reconstruction could not achieve sufficient control of pivot-shift when accompanying tibial tunnel coalition.

PURPOSE: To evaluate the effect of tibial tunnel coalition on knee rotatory laxity and clinical outcomes after double-bundle (DB) anterior cruciate ligament (ACL) reconstruction. METHODS: Forty-one patients who underwent anatomic DB ACL reconstruction were included prospectively. Three-dimensional computed tomography of the knee joint was obtained at approximately 1 year postoperatively to determine if tunnel coalition occurred. After excluding seven cases of femoral tunnel coalition, two groups were established based on the existence of a tibial tunnel coalition. The pivot-shift test was quantitatively evaluated on the basis of tibial acceleration preoperatively and at 1 year postoperatively. Two subjective scores, the International Knee Documentation Committee (IKDC) subjective and Lysholm scores, were also collected. The pivot-shift measurement and subjective scores were compared between the ACL-reconstructed knees with and without tibial tunnel coalition. The independent t test, Pearson's chi-square test, and Student t tests were used in data analysis. RESULTS: Twenty-one knees had tibial tunnel coalition (group C), whereas 13 knees did not have tunnel coalition(group N). Pivot-shift was significantly diminished postoperatively in both groups on the basis of the clinical examination and quantitative evaluations (p < 0.05). However, there was a small but significant difference in tibial acceleration demonstrating larger pivot-shift in group C (1.0 ± 0.6 m/s2) than in group N (0.5 ± 0.3 m/s2, p < 0.05). No significant difference was observed in the IKDC subjective and Lysholm scores (both n.s.). CONCLUSION: When the tibial tunnel coalition occurs after DB ACL reconstruction, knee rotatory laxity may not be restored in ACL-reconstructed knees, as expected in those without tunnel coalition. It is recommended that two tibial tunnels should be created separately when performing DB-ACL reconstruction to achieve better control of rotatory knee laxity. LEVEL OF EVIDENCE: III.