Search

98 Background: P-PSMA-101 is an autologous CAR-T therapy targeting PSMA, with a high percentage of stem cell memory T cells (TSCM) associated with efficacy, safety, and bone homing (particularly relevant to prostate cancer). It is manufactured using a novel non-viral transposon system (piggyBac) that creates high TSCM products. Genes are inserted encoding a PSMA-targeted Centyrin CAR, iCasp9-based safety switch, and DHFR to purify CAR-T cells. P-PSMA-101 completely eliminated tumors in intractable murine models of prostate cancer, providing rationale for this phase 1 trial (NCT04249947). Methods: Patients with mCRPC treated with or not eligible for a CYP17 inhibitor or second-generation antiandrogen, and a taxane were enrolled. P-PSMA-101 was manufactured from apheresed T cells and administered IV following a standard 3-day cy/flu lymphodepletion regimen. Dose escalation from 0.25-15 x 106 cells/kg is planned. Results: As of September 30, 2021, P-PSMA-101 had been administered to 10 heavily pretreated patients (median 7 prior regimens; range 3-15). Single infusions of 0.25 (n=5) to 0.75 (n=5) x 106 cells/kg have been assessed, with dose escalation continuing. P-PSMA-101 cells were shown to expand in blood via qPCR assay, peaking 2-3 weeks after infusion, consistent with the high percentage of TSCM. Significant antitumor responses were seen in this preliminary data set. Declines in PSA were seen in 7 patients (>50% in 3 and >99% in 1). Of 4 patients who had pre- and post-treatment FDG and PSMA-PET imaging, 3 demonstrated marked to complete resolution of abnormal uptake at known metastatic disease sites, with concordance in bone and CT scans, and/or circulating tumor cells (CTC). In 1 case, post-treatment tumor biopsy demonstrated infiltration by P-PSMA-101 CAR-T cells and elimination of tumor cells (pathologic complete response). Safety was consistent with expectations for a CAR-T product. CRS was seen in 60% (10% Gr ≥3) of patients. DLT was seen in 1 patient with macrophage activation syndrome/uveitis, and was the only Gr ≥3 CRS event. Immune effector cell-associated neurotoxicity syndrome (ICANS) has not occurred. CRS marker elevations were modest (max IL-6: 642.6 pg/mL). The most common AEs were cytopenias, infections, and constitutional symptoms (Gr ≥3 60%, 10%, and 0%), as expected with lymphodepletion. Treatable related ocular AEs were noted in 3 patients. Conclusions: These results parallel preclinical findings that P-PSMA-101 can produce marked efficacy in mCRPC, and very low doses are highly efficacious, consistent with unique product attributes such as the TSCM phenotype and bone tropism. This is the first report demonstrating profound antitumor effects of a novel PSMA-directed CAR-T-cell platform with concordant biochemical, radiographic, and pathologic parameters, demonstrating that therapeutic benefit of unarmored CAR-T cells in a major solid tumor is possible. Clinical trial information: NCT04249947.

P-BCMA-101 and P-BCMA-ALLO1, autologous and allogeneic BCMA targeting CAR-T cell therapies respectively, are manufactured using a novel transposon-based system called piggyBac (PB). They comprise a high percentage of desirable stem cell memory T-cells associated with efficacy and safety. Single agent P-BCMA-101 data was previously reported showing marked efficacy and minimal toxicity (PRIME study), and exploratory combination cohorts have also been evaluated. P-BCMA-ALLO1 is expected to be first assessed in patients by the time of this presentation. Here we report results for P-BCMA-101 exploratory cohorts and P-BCMA-ALLO1. PRIME is a Phase 1/2 clinical trial to assess the safety and efficacy of P-BCMA-101 in patients with RRMM (≥ 3 prior lines, including a proteasome inhibitor (PI) and an immunomodulatory agent (IMiD), or double refractory) (NCT03288493). Patients undergo apheresis to harvest T cells and P-BCMA-101 is then manufactured. Patients then receive a 3-day cyclophosphamide (300 mg/m 2/day) / fludarabine (30 mg/m 2/day) lymphodepletion (LDC) regimen. Following 2 days of rest, patients received a single administration of P-BCMA-101 in escalating doses starting at 0.75 x 10 6 CAR-T cells/kg. Similar P-BCMA-101 dose escalation was performed in combination cohorts with rituximab (Rit) or lenalidomide (Len). During the study, use of a nanoplasmid (NP) vector in CAR-T manufacturing was implemented to improve transposition efficiency and cell quality. As of June 30 th, 2021, 90 unique patients have been treated with P-BCMA-101 in 5 dose levels of single agent P-BCMA-101, 3 dose levels of P-BCMA-101 with Rit and 2 dose levels of P-BCMA-101 with Len. The median age is 61 years and 66/37% were M/F. Patients were heavily pre-treated with a median of 6 prior regimens (range 3-18), 100% had received prior PI and IMiD, 74% daratumumab and 63% ASCT. 13 patients were treated in combination with Rit and 9 in combination with Len. Like prior reports of P-BCMA-101, the safety profile compares favorably to other CAR-T. No dose limiting toxicities (DLT) were observed in the single agent or the combination cohorts. The addition of Rit or Len did not increase toxicity. Grade 1-2 CRS was seen in 25% (no G3/4) of patients and ICANS in 7% (2% G3). Tocilizumab was used in 11% of patients. There were no treatment related deaths or unexpected/off-target toxicities. The most common treatment emergent adverse events were cytopenias, infections and constitutional symptoms (≥ G3 neutropenia 74%, thrombocytopenia 30%, anemia 35%), as expected with CAR-T and LDC. The overall response rate (ORR) with the Rit and Len combinations was 73% and 71% respectively. The favorable safety profile allowed outpatient treatment in 23 (25%) patients. Founded on the P-BCMA-101 data, an allogeneic CAR-T (P-BCMA-ALLO1) using a similar PB construct was created and demonstrated excellent preclinical safety and efficacy leading to the initiation of a Phase 1 clinical trial (NCT04960579). P-BCMA-ALLO1 CAR-T cells are generated from healthy donor T-cells using Cas-CLOVER to knockout Beta-2 microglobulin to prevent host vs graft rejection, TCRβchain for graft vs host disease prevention. Proprietary booster molecules are utilized to increase manufacturing yield and cell quality. The CAR binding domain is a novel anti-BCMA VCAR that produced superior efficacy in tumor models. These constructs contain the same PB transposon transgene cassette, DHFR for gene selection and an iCasp9-based safety switch, successfully assessed in the PRIME study. The primary objective of the P-BCMA-ALLO1 phase 1 trial is to assess the safety and maximum tolerated dose (MTD) based on DLT in RRMM patients who have received a PI, IMid and CD38 mAb. Secondary objectives will assess the anti-myeloma effect of P-BCMA-ALLO1 and biomarkers. The protocol is designed to treat 40 RRMM patients in a standard 3+3 dose escalation with a P-BCMA-ALLO1 starting dose of 0.75 X 10 6 cells/kg. Patients will receive P-BCMA-ALLO1 following LDC with cyclophosphamide (300 mg/m 2/day) / fludarabine (30 mg/m 2/day X 3 days. In conclusion, transposon generated autologous CAR-T cells demonstrate excellent clinical activity with a favorable toxicity profile in RRMM, can be safely combined with Len and Rit, and administered in the outpatient setting. Allogeneic CAR-T cells generated from this platform represent a further advance. Current data from both clinical studies will be presented. Disclosures Derman: Sanofi: Membership on an entity's Board of Directors or advisory committees. Deol: Kite, a Gilead Company: Consultancy. Ali: BMS: Research Funding; Aduro: Research Funding; Poseida: Research Funding; Aduro: Consultancy; Amgen: Consultancy; Sanofi: Consultancy; Oncopeptides: Consultancy; BMS: Consultancy; Janssen: Consultancy. Dholaria: Pfizer: Research Funding; MEI: Research Funding; Poseida: Research Funding; Takeda: Research Funding; Janssen: Research Funding; Angiocrine: Research Funding; Jazz: Speakers Bureau; Celgene: Speakers Bureau. Berdeja: Bioclinica: Consultancy; BMS: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Lilly: Research Funding; Abbvie: Research Funding; EMD Sorono: Research Funding; Takeda: Consultancy, Research Funding; Prothena: Consultancy; Servier: Consultancy; Janssen: Consultancy, Research Funding; Karyopharm: Consultancy; Vivolux: Research Funding; Cellularity: Research Funding; Novartis: Research Funding; Poseida: Research Funding; CURIS: Research Funding; CRISPR Therapeutics: Consultancy, Research Funding; Acetylon: Research Funding; Amgen: Consultancy, Research Funding; Teva: Research Funding; Legend: Consultancy; Kite Pharma: Consultancy; Bluebird: Research Funding; Constellation: Research Funding; Glenmark: Research Funding; Genentech: Research Funding; Kesios: Research Funding. Cohen: GlaxoSmithKline: Consultancy, Research Funding; AstraZeneca: Consultancy; Genentech/Roche: Consultancy; BMS/Celgene: Consultancy; Novartis: Research Funding; Oncopeptides: Consultancy; Takeda: Consultancy; Janssen: Consultancy. Patel: Oncopeptides: Consultancy; BMS Celgene: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; Pfizer: Consultancy. Siegel: Karyopharm: Honoraria; Amgen Inc.: Honoraria; Takeda: Honoraria; Bristol Myers Squibb: Honoraria, Speakers Bureau; GlaxoSmithKline: Honoraria, Speakers Bureau; Celularity: Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Speakers Bureau. Nath: Actinium: Consultancy, Honoraria; Incyte: Consultancy, Honoraria. McArthur: Poseida: Current Employment, Current equity holder in publicly-traded company. McCaigue: Poseida: Current Employment, Current equity holder in publicly-traded company. Martin: Poseida: Current Employment, Current equity holder in publicly-traded company. Ghoddusi: Poseida: Current Employment, Current equity holder in publicly-traded company. Namini: Poseida: Current Employment, Current equity holder in publicly-traded company. Ostertag: Poseida: Current Employment, Current equity holder in publicly-traded company. Spear: Poseida: Current Employment, Current equity holder in publicly-traded company. Belani: Poseida Therapeutics: Current Employment, Current equity holder in publicly-traded company; Amgen: Current equity holder in publicly-traded company. Shah: Indapta Therapeutics: Consultancy; CareDx: Consultancy; BMS/Celgene: Research Funding; Amgen: Consultancy; Janssen: Research Funding; Bluebird Bio: Research Funding; Precision Biosciences: Research Funding; Karyopharm: Consultancy; Nektar: Research Funding; Oncopeptides: Consultancy; Poseida: Research Funding; Sanofi: Consultancy; CSL Behring: Consultancy; Kite: Consultancy; GSK: Consultancy; Sutro Biopharma: Research Funding; Teneobio: Research Funding.

Date Presented Accepted for AOTA INSPIRE 2021 but unable to be presented due to online event limitations. Idiopathic toe walking (ITW) causes significant physical impairment that affects a child’s occupational engagement. This research postulates a quasi-experimental study examining the responsiveness of a child with ITW using the Toe Tamer® protocol for 6 weeks during the COVID-19 pandemic. This research is relevant to the practice of OT by providing an individualized, accessible, cost-effective solution to ameliorate the effects of ITW in children. Primary Author and Speaker: M. Irma Alvarado Additional Authors and Speakers: Hannah Owens, Olivia Brand, and Sara Lieto Contributing Authors: Leigh Lehman, Ileana Seoane McCaigue

P-BCMA-101 is an autologous chimeric antigen receptor-T cell (CAR-T) therapeutic targeting BCMA and comprised of a high percentage of desirable stem cell memory T cells. P-BCMA-101 is manufactured using a novel transposon-based system called piggyBac and is designed to increase efficacy while minimizing toxicity. A phase 1/2, clinical trial is being conducted in patients with r/r MM (≥ 3 prior lines, including a proteasome inhibitor and an IMiD, or double refractory) to assess the safety and efficacy of P-BCMA-101 (NCT03288493). No pre-specified level of BCMA expression was required. Patients are apheresed to harvest T cells, P-BCMA-101 is then manufactured and administered to patients intravenously (IV) after a standard 3-day cyclophosphamide (300 mg/m2/day) / fludarabine (30 mg/m2/day) lymphodepletion regimen. As of 30Jun20, 43 patients had been treated with P-BCMA-101 (M/F 67%/33%, median age 60 years). Patients were heavily pre-treated (median of 7 prior regimens; range 3-18), with 100% having received proteasome inhibitors and IMiD, 93% daratumumab and 58% ASCT. This study was initially conducted as a dose escalation trial of single infusions of P-BCMA-101 from 0.75-15 x 106 cells/kg, preceded by standard lymphodepletion. Subsequently, exploratory cohorts with novel therapeutic strategies were evaluated. Using a modified manufacturing process, a median dose of 0.75 x 106 cells/kg were administered in cohorts including: P-BCMA-101 infusions in biweekly cycles; the addition of rituximab or lenalidomide pre- and post- lymphodepletion to prevent anti-CAR antibody development and increase T cell robustness, respectively; and single administration. The safety profile across the entire group was excellent for a CAR-T cell product which was attributed the gradual expansion of the Tscm cells (2-3 weeks to peak versus 3-7 days for most CAR-T cells). Cytokine release syndrome (CRS) was only seen in 17% of patients, with only one being grade 3 and one case of possible neurotoxicity reported (transient increase in confusion). Likewise, peak elevations of CRS markers were modest (maximum IL-6 level reported in any patient was 1631 pg/mL, orders of magnitude lower than levels frequently associated with severe CRS with CAR-T products). Only 3 patients required tocilizumab and no patients required ICU admission, safety switch activation or other aggressive measures. Based on the safety results the protocol was amended to allow fully outpatient CAR-T cell administration. There have been no patient deaths, DLTs or unexpected/off-target toxicities related to P-BCMA-101. The most common adverse events otherwise were cytopenias/infections and constitutional symptoms (≥ grade 3 neutropenia 79%, thrombocytopenia 30%, anemia 30%), as expected in CAR-T cell studies with lymphodepleting chemotherapy. Consistent with the high percentage of Tscm, circulating P-BCMA-101 cells were detected in blood by PCR, peaking at 2-3 weeks after infusion, and remaining detectable up to 1.5 years (ongoing at last follow-up). Response was seen to correlate with the Cmax and AUC of cell expansion, none of which correlated with dose administered. The overall response rate (ORR) for evaluable subjects (n=34) treated with single administration during the initial dose escalation was 57%. As there was not a definite dose response curve, but indications of better response at lower doses, additional cohorts were implemented focusing on the lower end of the dose range using product from the modified manufacturing process. Four patients were subsequently treated with cyclic administration, rituximab, lenalidomide or single administration at the lowest dose level with this manufacturing process (all treated with P-BCMA-101 within ~2 months prior to the data cut-off date), and thus far all have rapidly responded (100% ORR) and all responses are ongoing. The safety profile in these patients (including multiply infused patients) was no different than the overall population, with minimal CRS reported. In conclusion, current clinical data are consistent with preclinical findings that the novel design of P-BCMA-101 can produce significant efficacy, with remarkably low toxicity allowing for outpatient administration. Low doses appear highly efficacious and the modifications to manufacturing appear to have notably improved efficacy. Disclosures Costello: Poseida Therapeutics: Research Funding; Janssen: Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Cohen:Seattle Genetics: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Genentech/Roche: Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees; Kite Pharma: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees; Novartis: Other: Patents/Intellectual property licensed, Research Funding; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees; Takeda,: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees. Patel:Bristol Myers Squibb: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Precision Biosciences: Research Funding; Oncopeptides: Consultancy; Poseida: Research Funding; Nektar: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; Cellectis: Research Funding. Berdeja:Lilly: Research Funding; BMS: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Novartis: Research Funding; Bioclinica: Consultancy; Glenmark: Research Funding; Acetylon: Research Funding; Vivolux: Research Funding; Abbvie: Research Funding; CRISPR Therapeutics: Consultancy, Research Funding; CURIS: Research Funding; Janssen: Consultancy, Research Funding; Legend: Consultancy; Bluebird: Research Funding; Karyopharm: Consultancy; Kesios: Research Funding; Teva: Research Funding; Servier: Consultancy; Amgen: Consultancy, Research Funding; Cellularity: Research Funding; Celgene: Consultancy, Research Funding; Poseida: Research Funding; Prothena: Consultancy; Kite Pharma: Consultancy; EMD Sorono: Research Funding; Genentech, Inc.: Research Funding; Constellation: Research Funding. Shah:BMS, Janssen, Bluebird Bio, Sutro Biopharma, Teneobio, Poseida, Nektar: Research Funding; GSK, Amgen, Indapta Therapeutics, Sanofi, BMS, CareDx, Kite, Karyopharm: Consultancy. Ganguly:Kadmon: Other: Ad Board; KITE Pharma: Speakers Bureau; Settle Genetics: Speakers Bureau. Abedi:BMS, Gilead Sciences: Research Funding; AbbVie, BMS, Gilead Sciences, Seattle Genetics, Takeda: Speakers Bureau. Yalamanchili:Poseida Therapeutics: Current Employment, Current equity holder in private company. Gregory:Kesios: Research Funding; Sanofi: Research Funding; Janssen: Research Funding; Celularity: Research Funding; Teva: Research Funding; Vivolux: Research Funding; Lilly: Research Funding; Constellation: Research Funding; BMS: Research Funding; Celgene: Research Funding; Novartis: Research Funding; Poseida: Research Funding; CRISP Therapeutics: Research Funding; CURIS: Research Funding; Acetylon: Research Funding; Incyte Corporation: Consultancy; Bluebird: Research Funding; Amgen: Research Funding; AbbVie: Research Funding; Takeda: Research Funding; Genentech: Research Funding; Glenmark: Research Funding; EMD Sorono: Research Funding.

INTRODUCTION An exaggerated proinflammatory response to endotoxaemia can occur in obstructive jaundice. The aims of this study were to determine the hepatic proinflammatory and anti-inflammatory cytokine response to endotoxaemia in experimental biliary obstruction and to determine the source of interleukin-6 (IL-6) using immunohistochemistry. METHODOLOGY Male Wistar rats were randomized into three groups: bile duct ligation (BDL), sham operation, and control groups. They were weighed before surgery and after 1 week. On day 8, hepatic perfusion was performed with endotoxin (Escherichia coli 0111:B4). Serial samples of blood, effluent, and influent perfusate were analyzed for proinflammatory and anti-inflammatory cytokines. Ultrastructural assessment of sections of the liver was performed. RESULTS BDL animals lost weight in the first week compared with the sham and the control animals that gained weight. Liver function tests were elevated in the BDL group. Effluent biochemistry did not reveal liver injury as a result of perfusion. Ultrastructurally, there was no evidence of liver injury, with only active Kupffer cells, preservation of liver architecture, and minimal liver injury being detected. Serum tumor necrosis factor-α was not detected in any group before perfusion; however, serum IL-6 was higher in the BDL group. Portal endotoxaemia resulted in an increase in tumor necrosis factor-α, IL-6, and IL-10 in the BDL group. Fluorescence immunohistochemistry demonstrated IL-6 in the sinusoidal spaces and the cytoplasm of Kupffer cells. CONCLUSION There is an exaggerated proinflammatory and anti-inflammatory cytokine response to portal endotoxaemia in animals with jaundice compared with the sham group.

Orthopaedic tissue engineering combines the application of scaffold materials, cells and the release of growth factors. It has been described as the science of persuading the body to reconstitute or repair tissues that have failed to regenerate or heal spontaneously. In the case of bone regeneration 3-D scaffolds are used as a framework to guide tissue regeneration. Mesenchymal cells obtained from the patient via biopsy are grown on biomaterials in vitro and then implanted at a desired site in the patient's body. Medical implants that encourage natural tissue regeneration are generally considered more desirable than metallic implants that may need to be removed by subsequent intervention. Numerous polymeric materials, from natural and artificial sources, are under investigation as substitutes for skeletal elements such as cartilage and bone. For bone regeneration, cells (obtained mainly from bone marrow aspirate or as primary cell outgrowths from bone biopsies) can be combined with biodegradable polymeric materials and/or ceramics and absorbed growth factors so that osteoinduction is facilitated together with osteoconduction; through the creation of bioactive rather than bioinert scaffold constructs. Relatively rapid biodegradation enables advantageous filling with natural tissue while loss of polymer strength before mass is disadvantageous. Innovative solutions are required to address this and other issues such as the biocompatibility of material surfaces and the use of appropriate scaffold topography and porosity to influence bone cell gene expression.

Background Complex limb trauma often involves combined arterial and venous injury, and the resultant ischaemia–reperfusion injury (IRI) causes both local and remote organ injury. This study assessed the influence of the timing of restoration of venous drainage on IRI. Methods Male New Zealand white rabbits (n = 36) were randomized into six groups: sham operation (group 1) and unilateral hind limb arterial and venous occlusion for 1 h followed by no reflow for 2 h (group 2), arterial and venous reflow for 2 h (group 3), arterial reflow alone for 2 h (group 4), arterial reflow alone for 1 h followed by arterial and venous (delayed) reflow for a further 1 h (group 5), and pretreatment with an enteral combination antioxidant before occlusion of both artery and vein and delayed venous reflow (group 6). Plasma hydroperoxide (HPO) and glutathione peroxidase concentration, hind limb skeletal muscle and lung tissue wet: dry weight ratios and myeloperoxidase (MPO) concentration were measured. Results The plasma HPO level in the femoral vein effluent was significantly greater after delayed venous reflow (mean(s.e.m.) 2·02(0·54) μmol/l) than in control animals (0·98(0·10) μmol/l) (P < 0·05). There was also a significantly greater tissue wet: dry weight ratio after delayed venous reflow than in controls, in skeletal muscle (mean(s.e.m.) 6·89(0·14) versus 5·34(0·54); P < 0·05) and lung (9·20(1·14) versus 7·23(0·38); P < 0·05) tissue. Lung tissue MPO activity was significantly greater after delayed venous reflow compared with controls (3·20(0·28) versus 1·86(0·14) units/g; P < 0·005), and also in comparison to simultaneous arterial and venous reflow (2·40(0·24) units/g; P < 0·05). In the antioxidant pretreatment group there was no significant increase in plasma HPO concentration, tissue MPO level or tissue wet: dry weight ratio compared with the control group. Conclusion In combined major arterial and venous injury of the limb, delayed restoration of venous drainage leads to significantly greater local skeletal muscle injury and remote neutrophil-mediated lung injury. These results support the clinical rationale for early restoration not only of arterial inflow but also venous drainage by means of intraluminal shunts.

Background Sepsis and endotoxaemia occur frequently in biliary obstruction. Impaired Kupffer cell endocytosis is implicated in these events. Tumour necrosis factor and interleukin 6, secreted by Kupffer cells, are important mediators of sepsis. Kupffer cell clearance of endotoxin and secretion of cytokines in experimental obstructive jaundice were investigated. Methods Wistar rats were randomized to bile duct ligation, sham operation or control. Groups (n = 8) were studied 1 and 3 weeks after operation. Kupffer cell function was assessed using in situ hepatic perfusion. Results Clearance of endotoxin was significantly depressed 1 week (median (interquartile range) 20·3 (10·5–27·1) per cent) and 3 weeks (22·1 (20·2–23·2) per cent) after bile duct ligation compared with that in respective sham animals (35·5 (29·9–41·6) and 40·9 (37·7–47·0) per cent) and controls (39·5 (37·3–46·8) per cent). Secretion of tumour necrosis factor was significantly greater 1 week (1113·7 (706·5–1436·8) pg/ml) and 3 weeks (1118·2 (775·7–1484·1) pg/ml) following bile duct ligation compared with that in respective sham animals (114·3 (0–178·5) and 107·6 (63·7–166·4) pg/ml) and controls (0 (0–20·7) pg/ml). Interleukin 6 was not secreted by sham or control animals but was present in the perfusate from jaundiced animals at 1 and 3 weeks (52·5 (9·9–89·5) and 66·2 (60·2–193·1) pg/ml). Conclusion These data demonstrate simultaneous impairment of Kupffer cell clearance of endotoxin and increased secretion of proinflammatory cytokines in experimental obstructive jaundice. These diverse responses may contribute to the development of sepsis-related complications in biliary obstruction.

Background It has been suggested that bowel permeability is altered following abdominal aortic aneurysm surgery. The effect of ischaemia-reperfusion injury to the lower limb on the morphological structure, neutrophil infiltration and permeability of the bowel was investigated. Methods Histological assessment of the bowel was undertaken in five groups of Wistar rats: control, 3 h of bilateral hind limb ischaemia and 3 h of bilateral hind limb ischaemia followed by 1, 2 or 3 h of reperfusion. Using an everted gut sac model and 14 C-labelled polyethylene glycol, the effect of ischaemia-reperfusion on small bowel permeability was studied. Results The small bowel showed a significant decrease in mucosal thickness, villus height and crypt depth in animals subjected to ischaemia followed by 2-hr reperfusion (mean(s.e.m.) 420(15), 217(9) and 163(6) μm respectively) compared with controls (481(11), 245(6) and 195(6) μm) (P

In 30 patients undergoing elective repair of abdominal aortic aneurysm the intramucosal pH (pHi) of the sigmoid colon was measured. Blood for endotoxin assay was taken at intervals before, during and after surgery. Daily measurements were made of liver transaminase activity and of arterial partial pressure of oxygen (PaO2). The mean(s.e.m.) peak systemic endotoxin concentration in those who developed intramucosal acidosis (pHi below 7·00) was 90(14) pg/ml, compared with 42(5) pg/ml in those who did not (P < 0·01). In the 14 patients whose pHi fell below 7·00, the mean(s.e.m.) postoperative rise in aspartate transaminase activity was 346(74) per cent, compared with 181(20) per cent in those whose pHi remained above this level (P < 0·05). The mean(s.e.m.) postoperative ratio of PaO2, to the fraction of inspired oxygen was 177(11) mmHg in those with intramucosal acidosis, compared with 260(24) mmHg in those whose pHi remained above 7·00 (P < 0·01). These results demonstrate a relationship between bowel ischaemia, endotoxaemia and organ impairment following elective aneurysm repair.

Intestinal goblet cell numbers in two regions of the small intestine (20-30% and 60-70% distance form the pylorus) of male, 6- to 8-week-old C57 mice have been monitored following a 5-cysticercoid infection of the rat tapeworm, Hymenolepis diminuta. Test and sham-infected control mice were autopsied 0, 4, 8, 10, 14, and 28 days postprimary infection (p-1 degree-i) and 2, 4, 5, 7, and 14 days postsecondary infection (p-2 degree-i), administered 28 days p-1 degree-i. Results show a statistically significant increase in the number of mucus-containing goblet cells in both regions of the intestine during primary and secondary infections. Peak goblet cell numbers occurred on Day 8 p-1 degree-i and Day 5 p-2 degree-i in the 20-30% region and on Day 10 p-1 degree-i and Day 5 p-2 degree-i in the 60-70% region. In both regions, cell numbers declined to control levels by Day 14 p-1 degree-i, but remained significantly above control values 14 days p-2 degree-i. The increase in cell numbers correlated with an increase in goblet cell theca size and observable amounts of luminal mucus. The same infection regime in mice treated with cortisone elicited no goblet cell response. Male Wistar rats given a 10-cysticercoid infection and autopsied on Day 0, Day 10, and 15 months p-i showed a statistically significant increase in mucus-containing goblet cells only in the 60-70% region of intestine 10 days p-i; however, the worm burden was not eliminated. The functional significance of these results is discussed in relation to host immunity and murine cestode rejection.

A survey of the incidence of monocyte esterase deficiency in 4000 inpatients (including 808 with malignant neoplastic disease) and 474 normal controls was performed using an automated esterase method. A highly significant excess of patients with malignant disease and the deficiency was evident when compared with normal controls or all other patients. Within the group of patients with malignant disease the demonstrable excess occurred in B chronic lymphocytic leukaemia, non-Hodgkin's and Hodgkin's lymphoma, and carcinoma of the gastrointestinal tract. There was also a significant excess of patients with the deficiency attending the renal unit, both among patients who had had renal transplants and those who had not. A familial incidence of monocyte esterase deficiency was found in 19 (35%) of first degree relatives of those patients in whom family studies were done. It is suggested that the reason for the increased prevalence of the anomaly in these disorders might be that the diminution of esterase activity has a role in their development.

Orthopaedic tissue engineering combines the application of scaffold materials, cells and the release of growth factors. It has been described as the science of persuading the body to reconstitute or repair tissues that have failed to regenerate or heal spontaneously. In the case of bone regeneration 3-D scaffolds are used as a framework to guide tissue regeneration. Mesenchymal cells obtained from the patient via biopsy are grown on biomaterials in vitro and then implanted at a desired site in the patient's body. Medical implants that encourage natural tissue regeneration are generally considered more desirable than metallic implants that may need to be removed by subsequent intervention. Numerous polymeric materials, from natural and artificial sources, are under investigation as substitutes for skeletal elements such as cartilage and bone. For bone regeneration, cells (obtained mainly from bone marrow aspirate or as primary cell outgrowths from bone biopsies) can be combined with biodegradable polymeric materials and/or ceramics and absorbed growth factors so that osteoinduction is facilitated together with osteoconduction; through the creation of bioactive rather than bioinert scaffold constructs. Relatively rapid biodegradation enables advantageous filling with natural tissue while loss of polymer strength before mass is disadvantageous. Innovative solutions are required to address this and other issues such as the biocompatibility of material surfaces and the use of appropriate scaffold topography and porosity to influence bone cell gene expression.

To investigate the immunosuppressive properties of MSCs, in the present study we examined the immunogenicity of undifferentiated and trilineage-differentiated (chondrocytes, osteoblasts, and adipocytes) rat bone marrow-derived MSCs under xenogeneic conditions. After chondrogenic differentiation, rat bone marrow-derived MSCs stimulated human dendritic cells (hDCs) derived from peripheral blood monocytes, leading to eight- and fourfold higher lymphocyte proliferation and cytotoxicity than that of undifferentiated MSCs. The chondrogenic-differentiated MSCs were chemotactic to hDCs in Dunn chamber chemotaxis system and were rosetted by hDCs in rosette assays. Flow cytometry analysis revealed that chondrogenic-differentiated MSCs had promoted hDC maturation, causing higher CD83 expression in hDCs, whereas undifferentiated MSCs and osteogenic- and adipogenic-differentiated MSCs showed an inhibitory effect on hDC maturation. The costimulatory B7 molecules were upregulated only in the chondrogenic-differentiated MSCs. After blocking B7 molecules with specific monoclonal antibodies in the chondrogenic-differentiated MSCs, CD83 expression of cocultured hDCs was greatly reduced. In conclusion, chondrogenic differentiation may increase the immunogenicity of MSCs, leading to stimulation of dendritic cells. The upregulated expression of B7 molecules on the chondrogenic-differentiated MSCs may be partially responsible for this event.

Supplementation of mesenchymal stem cells (MSCs) during hematopoietic stem cell (HSC) transplantation alleviates complications such as graft-versus-host disease, leading to a speedy recovery of hematopoiesis. To meet this clinical demand, a fast MSC expansion method is required. In the present study, we examined the feasibility of using a rotary bioreactor system to expand MSCs from isolated bone marrow mononuclear cells. The cells were cultured in a rotary bioreactor with Myelocult medium containing a combination of supplementary factors, including stem cell factor and interleukin-3 and -6. After 8 days of culture, total cell numbers, Stro-1(+)CD44(+)CD34(-) MSCs, and CD34(+)CD44(+)Stro-1(-) HSCs were increased 9-, 29-, and 8-fold, respectively. Colony-forming efficiency-fibroblast per day of the bioreactor-treated cells was 1.44-fold higher than that of the cells without bioreactor treatment. The bioreactor-expanded MSCs showed expression of primitive MSC markers endoglin (SH2) and vimentin, whereas markers associated with lineage differentiation, including osteocalcin (osteogenesis), type II collagen (chondrogenesis), and C/EBP-alpha (CCAAT/enhancer-binding protein-alpha) (adipogenesis), were not detected. Upon induction, the bioreactor-expanded MSCs were able to differentiate into osteoblasts, chondrocytes, and adipocytes. We conclude that the rotary bioreactor with the modified Myelocult medium reported in this study may be used to rapidly expand MSCs.