Comparison of guidelines for biological ancillary materials used for the manufacture of gene and cellular therapy products in Asia.

BACKGROUND AIMS: Although biologiocal ancillay materials (AMs) have specific risk associated with their derivations, it plays key role to manufature cell and gene therapy (CGT) products. It is important to understand the regulation relevant to AMs for developers. METHODS: The authors investigated the guidelines and pharmacopeia (hereinafter referred to as "guidelines") for biological AMs used for the manufacture of CGT products in Asia (China, India, Japan, Korea and Taiwan). In addition, the authors benchmarked the relevant guidelines in the United States (US) and European Union (EU). RESULTS AND DISCUSSIONS: The guidelines could be classified into two types based on whether specific AMs are scoped: (i) general guidelines for risk assessment of AMs and (ii) guidelines for specific AMs. The authors compared the risk categories for each type of AM provided in the general guidelines between the US and China and the specific requirements for bovine serum and trypsin in the guidelines of China, Japan, Taiwan, US and EU. The authors further compiled in-depth descriptions of the respective regulations in China, India, Japan, Korea and Taiwan. There is limited availability of some guidelines for specific AMs. Moreover, there are no common requirements established across the surveyed countries and regions. Therefore, the authors suggest a risk assessment approach for AMs with consideration of their biological origin and traceability, production steps applied and ability to control or remove AMs from the final medicinal product over the CGT manufacturing process.

Nonclinical and quality assessment of cell therapy products: Report on the 4th Asia Partnership Conference of Regenerative Medicine, April 15, 2021.

The 4th Asia Partnership Conference of Regenerative Medicine (APACRM) was held online on April 15, 2021, to promote regulatory harmonization of regenerative medicine products throughout Asia. Recognizing domestic regulatory guidelines within each country and region, and their underpinning rationales, is an important initial step toward a convergence of regulations. The 4th APACRM consisted of an open dialog with regulatory agencies regarding nonclinical and quality settings for cell therapy products (CTPs) through industry presentations and panel discussions with regulatory agencies. The latest updates on regenerative medicine fields in each country and region, and specific regulatory schematics in Japan, were also introduced. The objective of this paper is to summarize the proceedings of the 4th APACRM for public dissemination and to foster further discussion in the future.

Inheritance and developmental pattern of cerebral hernia in the crested Polish chicken.

A number of crested chicken strains, such as the Polish chicken, exhibit a bony protuberance in the anterodorsal region of their skulls. The shape of their brain shows the anatomical peculiarity that is characterized by the upthrusting of cerebral hemispheres, called "cerebral hernia." Some early works suggested that this phenotype may be caused by a genetic factor and any modifiers influencing the development of brain and/or skull. However, the causative gene and its formation mechanism are still unclear. The present study is aimed to analyze the inheritance and ontogenic process of cerebral hernia in the crested Polish chicken. Firstly, we constructed the resource family with the Polish chicken and PNP inbred strain. Genetic analysis of this family revealed that cerebral hernia is controlled by a single autosomal recessive gene and is closely associated with crest formation. Furthermore, our morphological analysis of brain structures in the progenies suggested that the significant enlargement of brain cavity at later stages of embryos, particularly after 15 days of incubation, may be the main cause of cerebral hernia.

Immunolocalization of aquaporin-4 in the brain, kidney, skeletal muscle, and gastro-intestinal tract of chicken.

Aquaporin-4 (AQP4), a member of the water channel family, has high water permeability and multi-functional potentiality. Although an avian AQP4 homolog has recently been identified, its overall localization is still largely unknown. This study demonstrates the presence of AQP4 in several organs of chicken by using a specific chicken AQP4 antibody. Western blot analysis has revealed two bands of chicken AQP4 (30 and 32 kDa) in the brain, proventriculus, pectoral muscle, kidney, and ureter. The brain is the primary expression site of AQP4 in chicken. Immunohistochemical analysis of the brain has shown the highest AQP4 immunoreactivity around the cerebral ventricles, blood vessels, and the Purkinje cells. In peripheral organs, AQP4-immunoreactive elements have been observed in the ureter, glandular cells of the proventriculus, sarcolemma of the pectoral muscle, and the epithelium of the ceca and the rectum. Moreover, a heavily stained network of AQP4-immunoreactive fibers has been detected within the enteric plexuses.

Neonatal alloimmune thrombocytopenia caused by an antibody specific for a newly identified allele of human platelet antigen-7.

BACKGROUND: Neonatal alloimmune thrombocytopenia (NAIT) is a neonatal disorder characterized by maternal alloimmunization against fetal platelet (PLT) antigens inherited from the father. A healthy 30-year-old Japanese woman (Hit) gave birth to her second child after an uneventful pregnancy. Nine hours after birth, the infant presented with severe petechiae and a PLT count of 6 x 10(9)/L. STUDY DESIGN AND METHODS: To elucidate the maternal cause of NAIT in the infant, serologic and genetic studies, including PLT genotyping and sequence-based analysis, were conducted. Additionally, serologic screening for the new PLT antigen was performed. RESULTS: Serum from the NAIT infant's mother contained antibodies directed against a human PLT antigen (HPA) of the newborn. Using five-cell-lineage flow cytometry, we localized the antigen to a PLT glycoprotein (GP). Subsequent monoclonal antibody immobilization of PLT antigen assay and PLT immunofluorescence inhibition experiments localized the antigen to the GPIIIa subunit of the GPIIb/IIIa complex. GPIIIa localization was confirmed by sequence-based typing studies, which identified a 1297C>T (407proline>serine substitution) mutation on the ninth exon of the GPIIIa gene. This mutation identified the third allele of HPA-7. Anti-Hit(a) reacted with mutated GPIIIa-transfected cells but not with stable transfectants expressing wild-type GPIIIa. Serologic screening for Hit(a) in the Japanese population revealed a phenotypic frequency of approximately 0.0015. CONCLUSIONS: We identified a new third allele of HPA-7, which is characterized by a 1297C>T mutation in the GPIIIa gene. This 1297C>T allele was found in 0.15% of the Japanese population. An antibody against this antigen could be the cause of severe NAIT.

Polymerized structure and improved physical and biochemical material properties of carboxymethyl dextran-conjugated collagen.

Pepsin-solubilized collagen (PSC) was conjugated with carboxymethyl dextran (CMD) using cyanogen bromide to obtain a PSC-CMD film having improved physical properties, physiological properties, and cell affinity. The conjugation was confirmed by the loss of the alpha- and beta-subunit chains and the polymerized band on SDS-PAGE, and by a decrease in the isoelectric point to 3.2. PSC-CMD had a large polymerized structure with the 6 PSC and 228 CMD molecules. PSC-CMD was readily soluble in water, reconstructed a matrix with a less-ordered structure and a characteristic morphological shape, and lost platelet aggregation-inducing ability. The PSC-CMD film, cross-linked by ultraviolet irradiation, exhibited reduced solubility, moderate water vapor permeability, and increased flexibility. PSC-CMD coatings exhibited good cell attachment and growth for fibroblasts and vein endothrical cells.

Soft textural and emulsifiable gelatin formed by conjugating with fatty-acylated saccharide.

Ossein gelatin (OG) and low-molecular-weight ossein gelatin (LOG) were conjugated with a glucose/fructose stearic acid monoester (GE/FE), which had been prepared from the hydrolysate of a sucrose stearic acid diester, by the Maillard reaction to improve their physical properties. The molar ratio of each conjugate (GE/FE-OG or GE/FE-LOG) was about 1:1, and the conjugation resulted in a decreased isoelectric point. The GE/FE-OG gel showed a lower storage modulus, melting temperature, and enthalpy change than those for the OG gel at both the early and late stages of gelation, and each gel at pH 3.0 showed somewhat lower characteristics than those at pH 7.0. The conjugates acquired superior emulsifying ability, GE/FE-LOG in particular exhibiting markedly higher emulsifying ability in the acidic pH range, in the presence of NaCl, and over a wide temperature range. It is thus expected that conjugation with GE/FE could be effective for providing a new type of gelatin with a soft texture, easy melting, and emulsifying ability.

Discrimination of ionic pollutants except condensation nuclei of acid fog using an ultrasonic humidifier.

Fog droplets in the atmosphere are first produced by the activation of cloud condensation nuclei (CCN), which are originally some ionic compound. Subsequently, the nuclei grow by vapor diffusion. Fog droplets are polluted through the activation process and successive diffusion growth and residence (post activation). We cannot distinguish the effects of the two pollution processes of natural fog water samples. We found that fog droplets can be produced artificially without CCN using an ultrasonic humidifier. Because the artificial fog droplets are not polluted by CCN, the movement of the fog droplets in natural air will take up some pollutants in the air. Consequently, the two pollution processes of fog (the activation of CCN and the post activation process) can be discriminated using data from field experiments. This sampling analytical method is extremely important for further research regarding fog, clouds and environmental chemistry.

Improved physical and biochemical features of a collagen membrane by conjugating with soluble egg shell membrane protein.

Soluble egg shell membrane protein (SEP) prepared from the egg shell membrane by performic acid oxidation and pepsin digestion was applied to prepare a membrane conjugated with pepsin-solubilized collagen (PSC) by cross-linking with water-soluble carbodiimide. SEP enabled a conjugated membrane to be constructed with a well-developed network structure having thicker collagen fibrils and higher stability than the PSC membrane alone, as indicated by reduced solubility and collagenase-digestion, increased denaturation temperature, and superior flexibility. The SEP-PSC membrane (SEP:PSC of 1:1, w/w) had lower sweatiness than the PSC membrane. Fibroblasts and keratinocytes adhered to the SEP-PSC-coated wells (SEP:PSC of 1:10, w/w) as a fundamental model of an artificial skin similar to that to the PSC-coated wells due to similar expression level of several cell-adhesion proteins. An SEP-PSC coating accelerated the cell growth of fibroblasts, whereas the cell growth of keratinocytes did not change, or decreased slightly due to more rapid differentiation indicated by a more substantial expression of differentiation marker proteins than a PSC-coating without SEP.

Effect of long-range transport of air mass on the ionic components in radiation fog in northern Japan.

Water samples from radiation and upslope types of fog were collected at Takanosu Basin and Hachimantai mountain range of Akita Prefecture in northern Japan, respectively. The effect of the long-range transport of pollutants from the Asian Continent to the basin and the mountain range on the chemical characteristics of two types of fog was studied using chemical analysis data for the water samples as well as the back-trajectories of the air mass. In particular, non-sea-salt (nss)-SO4(2-) of radiation fog provided a much higher concentration than that of upslope fog, which suggests that nss-SO4(2-) might be transported mainly from the Asian Continent. The transport and uptake mechanism of pollutants in the radiation fog water were explained based on their accumulation in the basin due to anticyclonic subsidence and by the long-lasting fog behavior.