Towards gene therapy for IPEX syndrome.

Immune dysregulation polyendocrinopathy enteropathy X linked (IPEX) syndrome is an uncurable disease of the immune system, with immune dysregulation that is caused by mutations in FOXP3. Current treatment options, such as pharmacological immune suppression and allogeneic hematopoietic stem cell transplantation, have been beneficial but present limitations, and their life-long consequences are ill-defined. Other similar blood monogenic diseases have been successfully treated using gene transfer in autologous patient cells, thus providing an effective and less invasive therapeutic. Development of gene therapy for patients with IPEX is particularly challenging because successful strategies must restore the complex expression profile of the transcription factor FOXP3, ensuring it is tightly regulated and its cell subset-specific roles are maintained. This review summarizes current efforts toward achieving gene therapy to treat immune dysregulation in IPEX patients.

Chronologic Analysis of Tear Dynamics on Blinking Using Quantitative Manometry in Healthy Humans.

PURPOSE: To analyze the tear dynamics during blinking by measuring the inner pressure of the upper lacrimal drainage system. METHODS: This observational study involved 11 healthy bi- or tricenarian volunteers. Direct manometry was performed using a fiber optic pressure sensor inserted into the conjunctival sac, upper/lower canaliculus (5 mm from punctum), and inferior lacrimal sac (15 mm from punctum) during both involuntary and intentional tight blinking. Pressure was measured 200 times/second during 3 separate blinks and then chronologically analyzed. RESULTS: In all subjects of all locations during both types of blinking, the inner pressures during the stationary eyelid closing/opening were positive/approximately zero, while a positive/negative pressure spike was observed during the eyelid closing/opening movement. The averages of the maximum pressure in the spike during the intentional tight blinking (tPmax: mm Hg) in the conjunctival sac, upper/lower canaliculus, and lacrimal sac were 8.00, 12.39/12.93, and 10.59, respectively, while for the minimum (tPmin: mm Hg), the pressures were -3.18, -3.91/-3.43, and -3.31, respectively. The tPmax and tPmin in the lacrimal duct were positively correlated with that of the conjunctival sac, which suggested synchronism of the drainage system. However, the tPmax in the canaliculus was significantly higher than that of the conjunctival sac, which suggested that tears do not flow from the conjunctival sac into the lacrimal duct during eyelid closure. CONCLUSIONS: The upper lacrimal drainage system functions as one united lumen in the lacrimal pump. The positive /negative pressure spike is essential for the lacrimal pump to efficiently eject/aspirate the tear from the lacrimal/conjunctival sac.

Adjustable Medial Epicanthoplasty Using a Rotational Flap for Epiblepharon Repair.

ABSTRACT: This report aimed to introduce the new adjustable rotational skin flap for epicanthoplasty in combination with traditional epiblepharon repair by the modified Hotz procedure. This retrospective study involved 25 consecutive patients with superficial punctate keratitis secondary to epiblepharon complicated with epicanthal fold who underwent the combined surgery between 2019 and 2020. The mean patient age was 11.4 years in this study with a median follow-up was 8months (range, 3-12months). The rationale of the surgery was to release vertical tension of the eyelids by dissecting dense connective tissue beneath the epicanthal fold and to form a new medial canthus using a rotational skin flap supplied by the redundant the upper and/or lower eyelid skin excised during the epiblepharon repair. Postsurgical resolution ofsuperficial punctate keratitis and patient satisfaction was achieved in all patients. Additionally, there were no complications, and no revisional surgery was required in all patients for a median 8 months follow-up period. Utilizing a rotational skin flap during epiblepharon repair is a useful adjunct during epicanthoplasty surgery. This modification is well tolerated and allows for intraoperative adjustment, whereas minimizing scarring and allowing for improved tissue relaxation.

Activation of Mast-Cell-Derived Chymase in the Lacrimal Glands of Patients with IgG4-Related Ophthalmic Disease.

The purpose of this present study was to investigate the distribution and expression of chymase in the lacrimal glands (LGs) of patients afflicted with IgG4-related ophthalmic disease (IgG4-ROD). LGs from patients with severe canalicular obstruction were considered the control group. Toluidine blue staining confirmed a significant increase in the number of mast cells in the LGs obtained from the IgG4-ROD patients. In addition, immunostaining of serial sections from the LGs showed a significant increase in the number of chymase-positive cells and tryptase-positive cells in the IgG4-ROD LGs compared to the normal control LGs. The mRNA expression of chymase, tryptase, TGF-ß1, and collagen-I tended to increase in the IgG4-ROD LGs. Immunostaining of vimentin and α-smooth muscle actin (α-SMA) showed that myofibroblasts were the main cellular components in severely fibrotic regions of LGs in patients with IgG4-ROD. Linear regression analyses on the number of mast cells, chymase-positive cells, and tryptase-positive cells revealed significant positive correlations between those respective cells. Our findings suggest that chymase may play a role in the fibrotic disorder of IgG4-ROD LGs through the regulation of TGF-ß1 activation and collagen-I deposition, and that it may be a therapeutic target for patients afflicted with IgG4-ROD.

Pre-clinical development and molecular characterization of an engineered type 1 regulatory T-cell product suitable for immunotherapy.

BACKGROUND AIMS: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapeutic approach for many hematological disorders. However, allo-HSCT is frequently accompanied by a serious side effect: graft-versus-host disease (GVHD). The clinical use of allo-HSCT is limited by the inability of current immunosuppressive regimens to adequately control GvHD without impairing the graft-versus-leukemia effect (GvL) conferred by transplanted healthy immune cells. To address this, the authors have developed an engineered type 1 regulatory T-cell product called CD4IL-10 cells. CD4IL-10 cells are obtained through lentiviral transduction, which delivers the human IL10 gene into purified polyclonal CD4+ T cells. CD4IL-10 cells may provide an advantage over standard-of-care immunosuppressants because of the ability to suppress GvHD through continuous secretion of IL-10 and enhance the GvL effect in myeloid malignancies through targeted killing of malignant myeloid cells. METHODS: Here the authors established a production process aimed at current Good Manufacturing Practice (cGMP) production for CD4IL-10 cells. RESULTS: The authors demonstrated that the CD4IL-10 cell product maintains the suppressive and cytotoxic functions of previously described CD4IL-10 cells. In addition, RNA sequencing analysis of CD4IL-10 identified novel transcriptome changes, indicating that CD4IL-10 cells primarily upregulate cytotoxicity-related genes. These include four molecules with described roles in CD8+ T and natural killer cell-mediated cytotoxicity: CD244, KLRD1, KLRC1 and FASLG. Finally, it was shown that CD4IL-10 cells upregulate IL-22, which mediates wound healing and tissue repair, particularly in the gut. CONCLUSIONS: Collectively, these results pave the way toward clinical translation of the cGMP-optimized CD4IL-10 cell product and uncover new molecules that have a role in the clinical application of CD4IL-10 cells.

Negative impact of Interhospital Transfer on Clinical Outcomes of Mechanical Thrombectomy for Fast Progressive Stroke.

OBJECTIVES: The time-dependence of the clinical outcome of mechanical thrombectomy is higher in the "fast progressor" in whom cerebral ischemia progresses rapidly. The impact of time-consuming interhospital transfer (IT) on the clinical outcome of such patients is unknown. The effect on clinical outcomes of IT of fast progressors was investigated. METHODS: Among the patients enrolled in the Tokyo/Tama REgistry of Acute endovascular Thrombectomy, fast progressor cerebral ischemia cases were retrospectively investigated. In this study, a fast progressor was defined as a case with an Alberta Stroke Program Early CT Score less than 6 and last known well (LKW) to arterial puncture within 6 h. Patients' background characteristics, treatment progress, and the modified Rankin Scale (mRS) score at 3 months were examined. RESULTS: Of a total of 1182 patients, 92 (7.8%) were included, with 76 patients in the direct transfer (DT) group, and 16 patients in the IT group. Median LKW to reperfusion was 190 min and 272 min, respectively (P<.001). The number of patients with mRS scores 0-2 at three months was 22 (28.9%) in the DT group and 1 (6.2%) in the IT group. Interhospital transfer was an independent factor associated with worse outcomes (odds ratio 0.08, 95% confidence interval 0.01-0.87, P=.038). CONCLUSION: This study showed that, among fast progressor patients, the IT group had a worse prognosis than the DT group. To provide good clinical outcomes for fast progressor patients, those who are likely to undergo mechanical thrombectomy should be sent directly to a thrombectomy-capable center.

Phase-matching condition for THz wave generation via difference frequency generation using InxGa1-xSe mixed crystals.

Terahertz (THz) waves at 9.7, 10.1 and 10.6 THz were generated via difference frequency generation in high-quality InxG1-xaSe mixed crystals with a relatively high indium compositions (x = 0.040, 0.048, 0.074) grown from an indium flux. The phase-matching angle for THz wave generation was measured for each indium content. As a result, it is confirmed that the incident angle of the excitation light satisfying the phase-matching condition is shifted to a higher angle with an increase in the indium content.

Terahertz wave generation via difference frequency generation using 2D InxGa1-xSe crystal grown from indium flux.

We demonstrate the generation of THz waves (frequency 9.7 THz) using difference frequency generation in an InxGa1-xSe mixed crystal grown from In flux. The amount of indium and the lattice constant of the crystal were evaluated using electron micro probe analysis and X-ray diffraction, respectively. We believe that the Ga sites were substituted by In atoms in the InxGa1-xSe crystal because the In content, estimated according to the Vegard's law, was similar to that measured by EPMA. The maximum power of the generated THz wave was 39 pJ and the conversion efficiency was 1.7×10-5 J-1. This conversion efficiency was 28 times larger than that reported for undoped GaSe crystal.

Intracellular accumulation of PD-1 molecules in circulating T lymphocytes in advanced malignant melanoma: an implication for immune evasion mechanism.

BACKGROUND: The blockade of cell surface PD-1 ((sur)PD-1) by monoclonal antibodies, represented by nivolumab, provides the strategy to treat advanced malignant melanoma (AMM). The intracellular presence of PD-1 molecules have been reported in some T cell subsets, however, their kinetic association with those expressed on the cell surface, let alone their significance in antitumor immunity has been ill-investigated. METHODS: Intracellular PD-1 expression status in T cell subsets in AMM cases during nivolumab administration was chronologically characterized. The kinetics of PD-1 molecules within AMM-derived T cells was assessed in vitro in conjunction with their functional properties. RESULTS: Increase in (sur)PD-1 and intracellular PD-1 ((int)PD-1+) expression was characteristic for AMM T cells. After short-term culture, virtually (sur)PD-1- nivolumab-treated AMM T cells restore (sur)PD-1 expression, which could not be explained by the detachment of nivolumab from PD-1 epitopes alone. The blockade of trans-Golgi network resulted in the decrease in the extent of (sur)PD-1 recovery, suggesting the translocation of accumulated (int)PD-1 to the cell surface. Antigen-specific PD-1+ T cells significantly increased in (int)PD-1+ cells after treatment. In addition, a surge in (int)PD-1+CD4+ T cells was observed prior to the emergence of skin rash as an immune-related adverse event (irAE). CONCLUSIONS: Accumulated (int)PD-1 in T cells may contribute to enhanced immune evasion in AMM. Evaluation of intracellular PD-1 expression would be useful for better management of nivolumab-treated AMM patients in view of predicting treatment response and the incidence of irAE. Our findings further support the necessity of periodical administration of nivolumab for treating AMM.

Tregopathies: Monogenic diseases resulting in regulatory T-cell deficiency.

Monogenic diseases of the immune system, also known as inborn errors of immunity, are caused by single-gene mutations resulting in immune deficiency and dysregulation. More than 350 diseases have been described to date, and the number is rapidly expanding, with increasing availability of next-generation sequencing facilitating the diagnosis. The spectrum of immune dysregulation is wide, encompassing deficiencies in humoral, cellular, innate, and adaptive immunity; phagocytosis; and the complement system, which lead to autoinflammation and autoimmunity. Multiorgan autoimmunity is a dominant symptom when genetic mutations lead to defects in molecules essential for the development, survival, and/or function of regulatory T (Treg) cells. Studies of "Tregopathies" are providing critical mechanistic information on Treg cell biology, the role of Treg cell-associated molecules, and regulation of peripheral tolerance in human subjects. The pathogenic immune networks underlying these diseases need to be dissected to apply and develop immunomodulatory treatments and design curative treatments using cell and gene therapy. Here we review the pathogenetic mechanisms, clinical presentation, diagnosis, and current and future treatments of major known Tregopathies caused by mutations in FOXP3, CD25, cytotoxic T lymphocyte-associated antigen 4 (CTLA4), LPS-responsive and beige-like anchor protein (LRBA), and BTB domain and CNC homolog 2 (BACH2) and gain-of-function mutations in signal transducer and activator of transcription 3 (STAT3). We also discuss deficiencies in genes encoding STAT5b and IL-10 or IL-10 receptor as potential Tregopathies.

A novel method to assess the potential role of sweating abnormalities in the pathogenesis of atopic dermatitis.

Although atopic dry skin is believed to be caused by defects in skin genes important for maintaining skin barrier function, the role of sweat in atopic dermatitis (AD) has been apparently underestimated. Given the great capacity of sweat to maintain and increase skin hydration, defective sweating responses may be a logical place to look for changes that predispose individuals to the disease. We investigated how disease process and sweating defects progress from early asymptomatic stages to the onset of clinically apparent disease by employing the impression mould technique, which allows an accurate quantification of individual sweat gland/duct activity in relation to skin surface topography. Insensible and sensible sweating responses under baseline conditions and after thermal stimulus, respectively, were measured in various stages of AD patients and healthy controls. In controls, under baseline conditions, sweat ducts/glands at the dermal folds secreted basal levels of sweat (insensible sweating), thereby maintaining skin hydration. Not only such insensible sweating but also sensible sweating markedly decreased even in the earliest asymptomatic stage and the decrease was followed by compensatory hyperhidrosis at the ridge: leakage of sweat into the dermis could represent the initial event resulting in the decreased sweating and inflammation. The defects eventually progressed involving all of the ducts/glands to develop systemic dry skin. AD skin is characterized by varying degrees of functional impairment of sweat ducts/glands depending on the stage, and this defect would be among the reasons for the inability of AD patients to maintain skin hydration.

Association between Delirium and Prehospitalization Medication in Poststroke Patients.

PURPOSE: Medication is an important risk factor for delirium; however, the association between delirium and prehospitalization medication is unclear. We investigated the association between prestroke medication and poststroke delirium. MATERIALS AND METHODS: All patients hospitalized in the stroke care unit from September 2011 to September 2012 were selected, and their delirium symptoms, patient information, and pre- and poststroke medications were analyzed. Delirium was defined as a score of 4 or higher on the Intensive Care Delirium Screening Checklist. Factors that were related to delirium were extracted using univariate analysis, and the independent risk factors were determined using multivariate analysis. RESULTS: Of the 269 patients analyzed, 97 (36%) experienced delirium. Univariate analysis revealed significant differences between the delirium and nondelirium groups in age, dementia, previous cerebrovascular disease, craniotomy, all insertion-tube types, and 6 categories of prestroke medication. Prestroke polypharmacy was associated with poststroke delirium (P = .002). Multivariate analysis showed that taking antianxiety agents or sleep aids was an independent risk factor for delirium (odds ratio: 3.17, 95% confidence interval: 1.16-8.82). CONCLUSIONS: The present study suggests that prestroke medication affects the onset of poststroke delirium. These findings can contribute to the prediction and prevention of this condition.

Anti-BlyS antibody reduces the immune reaction against enzyme and enhances the efficacy of enzyme replacement therapy in Fabry disease model mice.

Formation of antibodies against a therapeutic enzyme is an important complication during enzyme replacement therapy (ERT) for lysosomal storage diseases. Fabry disease (FD) is caused by a deficiency of alpha-galactosidase (GLA), which results in the accumulation of globotriaosylceramide (GL-3). We have shown immune tolerance induction (ITI) during ERT in FD model mice by using an anti-B lymphocyte stimulator (anti-BlyS) antibody (belimumab). A single dose of the anti-BlyS antibody temporarily lowered the percentage of B cells and IgG antibody titer against recombinant human GLA. Administration of a low maintenance dose of the anti-BlyS antibody suppressed the B cell population and immunotolerance was induced in 20% of mice, but antibody formation could not be prevented. We then increased the maintenance dose of the anti-BlyS antibody and immunotolerance was induced in 50% of mice. Therapeutic enzyme distribution and clearance of GL-3 were also enhanced by a high maintenance dose of the anti-BlyS antibody.

Immunohistochemical dissection of cystic panfolliculoma focusing on the expression of multiple hair follicle lineage markers with an insight into the pathogenesis.

Panfolliculoma (PF) is a rare benign tumor with signs of differentiation toward all components of the hair follicle (HF). Cystic panfolliculoma (CPF) is a subset of PF with histological similarity to trichofolliculoma making the differential diagnosis difficult in some cases. Immunohistopathological investigations of PF have been reported; however, previous studies focused mostly on the expression profile of the outer root sheath markers. Herein, we report a case of CPF. A panel of antibodies was developed and used for the detection of the expression of cytokeratin (CK) 10, CK13, CK14, CK15, hair-hard keratin (AE13) and EpCAM within the lesion, supporting the differentiation of all epithelial lineages of the HF and the diagnosis of CPF. Immunohistopathological examinations clearly showed the spatial distribution pattern of each subset of tumor cells with distinct HF differentiation marker expression. Intriguingly, fibroblastic dermal cells were distributed preferentially near CK15-negative epithelium or CK13-positive HF-like structures, suggesting a role for epithelial-mesenchymal interactions (EMIs) in CPF pathogenesis. Further characterization of EMIs between the tumor and surrounding mesenchymal cells in CPF may provide an explanation for immature HF differentiation. These findings suggest that the more intense and coordinated EMI in the analogous tumorigenesis gives rise to mature HF structures, resulting in trichofolliculoma, which may explain their histological resemblance.

Clinical Evaluation of Implant-Supported Removable Partial Dentures With a Stress-Breaking Attachment.

PURPOSE: The stress-breaking ball (SBB) attachment can distribute the occlusal force equally between the alveolar ridge and the implants. The purpose of this study was to evaluate the implant-supported distal extension removable partial dentures (RPDs) with SBB attachment in 10 patients who were partially edentulous. METHODS: This randomized crossover study was designed to compare the function of RPDs with and without healing abutments and SBB attachments to support the posterior aspects of the RPDs. Mandibular jaw movements during mastication and the occlusal force and contact area were measured with a commercially available tracking device and pressure-sensitive sheets. Using a visual analog scale, 4 criteria-chewing, retention, stability, and comfort-were evaluated. All of the data obtained were analyzed using a 1-way analysis of variance (α = 0.05). RESULTS: There were no significant differences in either the mean time or the coefficient of variation among the SBB attachments and healing abutments of implant-supported removable partial dentures (ISRPDs) and conventional removable partial dentures (CRPDs). SBB attachments and healing abutments of ISRPDs had greater forces and contact areas than those of CRPDs with significant differences. For all criteria, patients preferred SBB attachments to healing abutments and CRPDs. CONCLUSIONS: The implant-supported distal extension RPDs with SBB attachment improved denture stability and patients' satisfaction.

An Adsorbate Discriminatory Gate Effect in a Flexible Porous Coordination Polymer for Selective Adsorption of CO2 over C2H2.

The adsorptive separation of C2H2 and CO2 via porous materials is nontrivial due to the close similarities of their boiling points and kinetic diameters. In this work, we describe a new flexible porous coordination polymer (PCP) [Mn(bdc)(dpe)] (H2bdc = 1,4-benzenedicarboxylic acid, dpe = 1,2-di(4-pyridyl)ethylene) having zero-dimensional pores, which shows an adsorbate discriminatory gate effect. The compound shows gate opening type abrupt adsorption for C2H2 but not for CO2, leading to an appreciable selective adsorption of CO2 over C2H2 at near ambient temperature (273 K). The origin of this unique selectivity, as unveiled by in situ adsorption-X-ray diffraction experiments and density functional theory calculations, is due to vastly different orientations between the phenylene ring of bdc and each gas in the nanopores. The structural change by photochemical transformation of this PCP via [2 + 2] photodimerization leads to the removal of inverse CO2/C2H2 selectivity, verifying the mechanism of the guest discriminatory gate effect.

Postsynthesis of h-BN/Graphene Heterostructures Inside a STEM.

Combinations of 2D materials with different physical properties can form heterostructures with modified electrical, mechanical, magnetic, and optical properties. The direct observation of a lateral heterostructure synthesis is reported by epitaxial in-plane graphene growth from the step-edge of hexagonal BN (h-BN) within a scanning transmission electron microscope chamber. Residual hydrocarbon in the chamber is the carbon source. The growth interface between h-BN and graphene is atomically identified as largely N-C bonds. This postgrowth method can form graphene nanoribbons connecting two h-BN domains with different twisting angles, as well as isolated carbon islands with arbitrary shapes embedded in the h-BN layer. The electronic properties of the vertically stacked h-BN/graphene heterostructures are investigated by electron energy-loss spectroscopy (EELS). Low-loss EELS analysis of the dielectric response suggests a robust coupling effect between the graphene and h-BN layers.

The production of human glucocerebrosidase in glyco-engineered Nicotiana benthamiana plants.

For the production of therapeutic proteins in plants, the presence of ß1,2-xylose and core α1,3-fucose on plants' N-glycan structures has been debated for their antigenic activity. In this study, RNA interference (RNAi) technology was used to down-regulate the endogenous N-acetylglucosaminyltransferase I (GNTI) expression in Nicotiana benthamiana. One glyco-engineered line (NbGNTI-RNAi) showed a strong reduction of plant-specific N-glycans, with the result that as much as 90.9% of the total N-glycans were of high-mannose type. Therefore, this NbGNTI-RNAi would be a promising system for the production of therapeutic glycoproteins in plants. The NbGNTI-RNAi plant was cross-pollinated with transgenic N. benthamiana expressing human glucocerebrosidase (GC). The recombinant GC, which has been used for enzyme replacement therapy in patients with Gaucher's disease, requires terminal mannose for its therapeutic efficacy. The N-glycan structures that were presented on all of the four occupied N-glycosylation sites of recombinant GC in NbGNTI-RNAi plants (GC(gnt1) ) showed that the majority (ranging from 73.3% up to 85.5%) of the N-glycans had mannose-type structures lacking potential immunogenic ß1,2-xylose and α1,3-fucose epitopes. Moreover, GC(gnt1) could be taken up into the macrophage cells via mannose receptors, and distributed and taken up into the liver and spleen, the target organs in the treatment of Gaucher's disease. Notably, the NbGNTI-RNAi line, producing GC, was stable and the NbGNTI-RNAi plants were viable and did not show any obvious phenotype. Therefore, it would provide a robust tool for the production of GC with customized N-glycan structures.

Pathological role of regulatory T cells in the initiation and maintenance of eczema herpeticum lesions.

It remains unknown why the occurrence of eczema herpeticum (EH) caused by an extensive disseminated cutaneous infection with HSV-1 or HSV-2 is associated with the exacerbation of atopic dermatitis lesions after withdrawal of treatment. Although regulatory T cells (Tregs) limit the magnitude of HSV-specific T cell responses in mice, their role in the induction and resolution of EH has not been defined. We initially investigated the frequencies, phenotype, and function of Tregs in the peripheral blood of atopic dermatitis with EH (ADEH) patients at onset and after clinical resolution, atopic dermatitis patients without EH, and healthy controls. Tregs with the skin-homing phenotype and the activated/induced phenotype were expanded at onset and contracted upon resolution. Treg-suppressive capacity was retained in ADEH patients and, the expanded Tregs suppressed IFN-γ production from HSV-1-specific CD8(+) and CD4(+) T cells. The increased frequency of CD14(dim)CD16(+) proinflammatory monocytes (pMOs) was also observed in the blood and EH skin lesions. Thus, pMOs detected in ADEH patients at onset were characterized by an increased ability to produce IL-10 and a decreased ability to produce proinflammatory cytokines, unlike their normal counterparts. Our coculture study using Tregs and pMOs showed that the pMOs can promote the expansion of inducible Tregs. Tregs were detected frequently in the vicinity of HSV-expressing and varicella zoster virus-expressing CD16(+) monocytes in the EH lesions. Expansions of functional Tregs, together with pMOs, initially required for ameliorating excessive inflammation occurring after withdrawal of topical corticosteroids could, in turn, contribute to the initiation and progression of HSV reactivation, resulting in the onset of EH.

Numerical and experimental investigations of human swimming motions.

This paper reviews unsteady flow conditions in human swimming and identifies the limitations and future potential of the current methods of analysing unsteady flow. The capability of computational fluid dynamics (CFD) has been extended from approaches assuming steady-state conditions to consideration of unsteady/transient conditions associated with the body motion of a swimmer. However, to predict hydrodynamic forces and the swimmer's potential speeds accurately, more robust and efficient numerical methods are necessary, coupled with validation procedures, requiring detailed experimental data reflecting local flow. Experimental data obtained by particle image velocimetry (PIV) in this area are limited, because at present observations are restricted to a two-dimensional 1.0 m(2) area, though this could be improved if the output range of the associated laser sheet increased. Simulations of human swimming are expected to improve competitive swimming, and our review has identified two important advances relating to understanding the flow conditions affecting performance in front crawl swimming: one is a mechanism for generating unsteady fluid forces, and the other is a theory relating to increased speed and efficiency.