Effect of Scarf and Chevron combined with Akin on postoperative balance in patients with moderate to severe foot bunion.

OBJECTIVE: The aim of this study was to analyze the effect of Scarf and Chevron combined with Akin on a postoperative balance of patients with moderate to severe foot bunion. PATIENTS AND METHODS: One hundred (100 feet) patients with moderate to severe bunion cysts treated at our hospital from January 2019 to January 2022 were retrospectively selected as subjects and divided into 2 groups according to their surgical procedure. The control group received Scarf combined with Akin, and the study group received Chevron combined with Akin. Oxidative stress mediators [late oxidized protein product (AOPP), lipid peroxide (LPO)], inflammatory factors [interleukin-1ß (IL-1ß), procalcitonin (PCT)], Hallux valgus angle (HVA), intermetatarsal angle (IMA), distal metatarsal joint angle (DMAA) Angle, ankle-hind foot American Orthotic Foot and Ankle Association (AOFAS) score, pain visual analog scale (VAS) score and balance Berg Balance Scale (BBS) score were compared between the two groups before and after surgery. The effectiveness and safety of the operation were compared. RESULTS: The levels of AOPP and LPO in the study group decreased most significantly, t=1.081 and 10.850, p=0.001; the levels of IL-1ß and PCT in the study group increased most significantly, t=16.970 and 12.260, p=0.001; the indexes of HVA, IMA, and DMAA in the study group increased significantly, t=11.890, 11.550, and 12.670, p=0.001; the AOFAS and BBS scores in the study group increased significantly, while the VAS score in the study group decreased significantly, t=14.760, 13.580, 5.994, p=0.001; the total effective rate of treatment in the study group was the highest, χ²=6.960, p=0.00; the total incidence of complications in the study group was the lowest, χ²=1.834, p=0.175. CONCLUSIONS: Chevron combined with Akin is more effective than Scarf combined with Akin in treating moderate to severe foot bunion, the former is more minimally invasive and has a better effect in promoting postoperative balance recovery.

Lactoferrin causes IgA and IgG2b isotype switching through betaglycan binding and activation of canonical TGF-ß signaling.

Lactoferrin (LF), a pleiotropic iron-binding glycoprotein, is known to modulate the humoral immune response. However, its exact role in Ig synthesis has yet to be elucidated. In this study, we investigated the effect of LF on Ig production by mouse B cells and its underlying mechanisms. LF, like transforming growth factor (TGF)-ß1, stimulated B cells to produce IgA and IgG2b, while downregulating other isotypes. Using limiting dilution analysis, LF was shown to increase the frequency of IgA-secreting B-cell clones. This was paralleled by an increase in Ig germ-line α (GLα) transcripts, indicating that LF plays a role as an IgA switch factor. Interestingly, LF directly interacted with betaglycan (TGF-ß receptor III, TßRIII) and in turn induced phosphorylation of TßRI and Smad3 through formation of the TßRIII/TßRII/TßRI complex, leading to IgA isotype switching. Peroral administration of LF increased intestinal/serum IgA production as well as number of IgA plasma cells in lamina propria. Finally, we found that LF has an adjuvant activity when nontoxigenic Salmonella typhimurium was inoculated perorally, conferring protection against intragastrical infection of toxigenic S. typhimurium. These results suggest that LF has an important effect on the mucosal/systemic IgA response and can contribute to protection against intestinal pathogens.

Molecular cloning and characterization of the STAT gene in Hyphantria cunea haemocytes.

A new insect member of the signal transducer and activator of transcription (STAT) family of transcription factors, Hyphantria cunea STAT (HcSTAT), was cloned from the lepidopteran H. cunea. The domain involved in DNA interaction and the Src homology 2 (SH2) domain were well conserved. During all developmental stages, the gene was expressed at a low level in the haemocytes, fat body cells, midgut, epidermis and Malpighian tubules. The haemocytes and Malpighian tubules showed transcriptional activation of HcSTAT upon Gram-negative and Gram-positive bacterial challenges. These challenges increased the induction and nuclear translocation of the HcSTAT protein that recognizes a STAT target site in H. cunea haemocytes. In vivo treatment with sodium orthovanadate translocated HcSTAT to the haemocyte nucleus. This study shows the involvement of the haemocyte Janus kinase/STAT pathway after microbial infection in lepidopteran insects.

Population genetic structure of the migratory rice leaf roller, Cnaphalocrocis medinalis (Lepidoptera: Pyralidae), inferred from the mitochondrial A+T-rich region and nuclear ITS2 sequences.

The population genetics of the migratory rice leaf roller, Cnaphalocrocis medinalis (Lepidoptera: Pyralidae), was characterized using the maternally inherited mitochondrial A+T-rich region and bi-parentally inherited nuclear internal transcribed spacer 2 (ITS2). One hundred and eighty-seven specimens of the rice leaf roller collected from 13 Korean and Chinese localities revealed 94 A+T-rich region haplotypes, ranging in sequence length from 339 to 348 bp and 129 ITS2 sequence types, ranging from 444 to 450 bp, with maximum sequence divergences of 4.55 and 4.43%, respectively. The finding of almost no significant F(ST), even among Chinese and Korean localities, except for one Chinese island population (ITS2 only), and the finding of genetic variance principally at the within-population level indicate the genetic structure characteristics of a migratory insect that is well connected among populations due to high gene flow. Detection of significant F(ST) estimates of one offshore island population in China (Haikou) compared to most others only by ITS2 rather than by the mitochondrial A+T-rich region, as well as the somewhat higher degree of genetic differentiation seen on ITS2, suggest the importance of female dispersal. Structural analysis of the A+T-rich region revealed a poly-T stretch (10-16 bp), a microsatellite-like AT repeat (10-14 repeats), and a 5-bp long-motif "ATTTA". The typical 5-bp long conserved motif sequence (ATAGA) previously detected in other lepidopterans was found to be ATAG in the C. medinalis A+T-rich region.

Modulation of MnSOD protein in response to different experimental stimulation in Hyphantria cunea.

A manganese superoxide dismutase (MnSOD) gene was cloned from the fall webworm, Hyphantria cunea. MnSOD cDNAs encode precursor proteins of 215 amino acid residues. H. cunea MnSOD possesses the metal binding ligands of 3 histidines and 1 aspartic acid common to MnSODs. The deduced amino acid sequences of the H. cunea MnSOD cDNA showed 76% identity to Bombyx mori MnSOD and 56-62% identity to MnSOD sequences from other species. MnSOD and copper/zinc superoxide dismutase (Cu/ZnSOD) is expressed in all tissues of H. cunea. MnSOD expression changed at a trace-level in infected larvae, while Cu/ZnSOD expression strongly changed against Gram-positive and Gram-negative bacteria, and fungi. Environmental stresses such as different artificial photoperiods (24L:0D), ultraviolet irradiation (312 nm), and starvation condition increased Cu/ZnSOD expression, MnSOD expression, on the other hand, was increased by starvation. Moreover, MnSOD and Cu/ZnSOD expression showed no significant change in the 0L:24D condition. MnSOD and Cu/ZnSOD expression in H. cunea also significantly increased at high (37°C) and low (4°C) temperature. Oxidative stress induced by 10% H(2)O(2) reduced the expression levels of MnSOD and Cu/ZnSOD. However, paraquat-induced oxidative stress reduced MnSOD expression but increased Cu/ZnSOD expression. These results suggest that Cu/ZnSOD may play a larger role than MnSOD as a superoxide anion scavenger against oxidative stress in H. cunea.

Comparative analysis of two attacin genes from Hyphantria cunea.

A full-length clone corresponding to attacin was isolated from a cDNA library made from fat body of immunized Hyphantria cunea larvae. This newly isolated attacin B shows characteristics different from those previously reported for attacin A. The two attacin cDNAs encode precursor proteins of 233 and 248 amino acid residues, respectively. The two attacins show 45.9% identity at the amino acid level, and 35.2% identity at the nucleotide level. Attacins A and B of H. cunea show significant identities with the attacins of Lepidoptera. Attacin B is a typical glycine-rich protein, while attacin A is leucine-rich. Attacin B is expressed from last instar larvae to adult, while attacin A showed stage-specific expression during the prepupal and pupal stages. Attacins A and B are predicted to have different secondary structure in that attacin A has no tendency to form helices but attacin B contains a substantial number of helices. Attacin A is induced at a trace level in infected larvae, while attacin B is strongly induced against Gram-positive and negative bacteria, fungi, and viruses. The attacin B transcripts were detected in fat body, epidermis and hemocytes after injection with Escherichia coli, Citrobacter freundii, or Candida albicans, but not in the midgut and Malpighian tubule. Recombinant attacin A showed no antibacterial activity, while recombinant attacin B showed strong antibacterial activity in proportion to the amount of the protein injected.

The mitochondrial genome of the Korean hairstreak, Coreana raphaelis (Lepidoptera: Lycaenidae).

We determined the complete nucleotide sequences of the mitochondrial genome (mitogenome) of the Korean hairstreak, Coreana raphaelis (Lepidoptera: Lycaenidae). The entire mitochondrial DNA (mtDNA) molecule was 15,314 bp long. The C. raphaelis genes were in the same order and orientation as the completely sequenced mitogenomes of other lepidopteran species, except for the presence of an extra copy of tRNA(Ser)(AGN). High similarity in primary sequence and secondary structure between the two tandemly located copies of the tRNA(Ser)(AGN) suggest a recent duplication of an original single tRNA(Ser)(AGN). The DHU arm of the two copies of tRNA(Ser)(AGN) formed a simple loop as seen in many other metazoan mt tRNA(Ser)(AGN). The putative initiation codon for the C. raphaelis COI gene appears to be a tetranucleotide, TTAG, found commonly in the sequenced lepidopterans. ATPase8, ATPase6, ND4L and ND6 genes, which are next to another protein-coding gene at their 3' end all had the sequences potential to form a hairpin structure, suggesting the importance of such a structure for precise cleavage of the mature protein-coding genes.

Baculovirus expression vectors that incorporate the foreign protein into viral occlusion bodies.

Current baculovirus expression systems typically produce soluble proteins that accumulate within the infected insect cell or are secreted into the growth medium. A system has now been developed for the incorporation of foreign proteins, along with the matrix protein, polyhedrin, into baculovirus occlusion bodies. Initial studies showed that a recombinant virus expressing a translational fusion between polyhedrin and GFP did not form occlusion bodies. However, a baculovirus coexpressing native polyhedrin and the polyhedrin-GFP fusion protein formed occlusion bodies that fluoresced under UV light, demonstrating that they included the polyhedrin-GFP fusion protein. This was confirmed by immunoblot analysis. Thus, incorporation of a foreign protein into occlusion bodies depends on an interaction between native polyhedrin and the polyhedrin fusion protein. Electron microscopy demonstrated that the occlusion bodies containing GFP also incorporated virions as expected. These ColorPol occlusion bodies were as infectious to insect larvae as occlusion bodies produced by wild-type virus. This new system expands the capabilities for foreign gene expression by baculoviruses, which has implications for biopesticide design, novel vaccine delivery systems, and fusion protein purification applications.

Molecular cloning and expression of a cDNA encoding the luciferase from the firefly, Pyrocoelia rufa.

To clone a cDNA encoding the luciferase of the firefly, Pyrocoelia rufa, we have constructed a cDNA library and isolated the luciferase gene using PCR with gene specific primers. Sequence analysis of the cDNA encoding the luciferase of P. rufa revealed that the 1647 bp cDNA has an open reading frame of 548 amino acid residues. The deduced amino acid sequences of the luciferase gene of P. rufa showed 98.9% homology to that of P. miyako. Phylogenetic analysis further confirmed the deduced amino acid sequences of the P. rufa luciferase gene belonged to the same subfamily, Lampyrinae. Southern blot analysis suggested possible presence of the P. rufa luciferase gene as a single copy and Northern blot analysis confirmed light organ-specific expression pattern at the transcriptional level. The cDNA encoding the luciferase of P. rufa was expressed as a 69 kDa band in baculovirus-infected insect cells and the recombinant baculovirus-infected cell extracts emitted luminescence in the luciferase activity assay.

Molecular cloning and characterization of a cDNA encoding a transferrin homolog from Bombyx mori.

We isolated a cDNA representing a message that was strongly induced by injection with E. coli in Bombyx mori. The 2160 bp cDNA has an open reading frame of 644 amino acids and the deduced product a predicted molecular mass of 71 kDa. The cDNA sequence shared high homology with the transferrins known so far, and its deduced peptide had unique features of transferrins, that is, sites of cystein residues and iron binding. We suggest that the B. mori transferrin plays an important role in the self-defense system.

The morphology of the polyhedra of a host range-expanded recombinant baculovirus and its parents.

The host range-expanded recombinant baculovirus, RecB-8 was isolated from BmN-4 cells coinfected with Autographa californica and Bombyx mori nuclear polyhedrosis viruses. Its genome was compared with those of its parents by restriction endonuclease digestion and their polyhedra compared in an electron microscope. Interestingly, the polyhedra of RecB-8 were tetrahedral although the polyhedrin gene was the same as that of the BmNPV parent which has icosahedral polyhedra. Thus the morphology of the RecB-8 polyhedra resulted from host cell factors and/or another viral genome in the host cells.

Isolation of a strain of Bacillus thuringiensis ssp. kurstaki HD-1 encoding delta-endotoxin Cry1E.

A strain of Bacillus thuringiensis, STB-1, toxic against Spodoptera exigua, was isolated. Bacillus thuringiensis STB-1 produced bipyramidal inclusions and reacted with the H antiserum of B. thuringiensis ssp. kurstaki. The plasmid and protein profiles of B. thuringiensis STB-1 were compared with those of its reference strains, ssp. kurstaki and ssp. kenyae. To verify the gene type of B. thuringiensis STB-1, PCR analysis was performed with Spodoptera-specific cry gene primers. The result showed that B. thuringiensis STB-1, unlike its reference strains, had crylAa, crylAb, crylAc and crylE, suggesting that B. thuringiensis STB-1 was a unique strain with respect to gene type. In addition, B. thuringiensis STB-1 showed a high level of toxicity against both S. exigua and Bombyx mori, whereas B. thuringiensis ssp. kurstaki HD-1 or ssp. kenyae showed a high level of toxicity against only Bombyx mori or S. exigua, respectively.

Effect of silkworm hemolymph on the expression of E. coli beta-galactosidase in insect cell lines infected with recombinant baculoviruses.

The effects of silkworm hemolymph on the expression of foreign genes by recombinant baculoviruses in cell lines were studied. The expression efficiency of beta-galactosidase by recombinant virus containing the E. coli lacZ gene at various concentrations of hemolymph and FBS was determined in BmN and Sf cell lines. The addition of hemolymph to the medium containing FBS accelerated the expression of beta-galactosidase by recombinant viruses in both cells. It was more effective in BmN cells than in Sf cells. Hemolymph was most effective in enhancing virus multiplicity under conditions of 5% FBS.

Characterization of recombinant Autographa californica nuclear polyhedrosis virus (NPV) expressing the beta-galactosidase gene in both Sf21 and Bm5 cells by Bombyx mori NPV p143 helicase gene.

Genomic DNA of recombinant AcNPV expressing beta-galactosidase was cotransfected with p143 helicase gene of BmNPV into Sf21 cells. Ac-Bm hybrid viruses capable of replicating in both Bm5 and Sf21 cells were isolated. Ac-Bm hybrid viruses expressing beta-galactosidase either at the highest (Ac-Bm hybrid virus-HE) or lowest (Ac-Bm hybrid virus-LE) level were chosen for the characterization of beta-galactosidase expression in Bm5 and Sf21 cells. Expression level of beta-galactosidase and replication of Ac-Bm hybrid virus-HE in Sf21 cells were nearly identical to those of recombinant AcNPV. Furthermore, replication of Ac-Bm hybrid virus-HE in Bm5 cells was similar to that of wild-type BmNPV, and Ac-Bm hybrid virus-HE clearly expressed beta-galactosidase in Bm5 cells. However, expression of beta-galactosidase by Ac-Bm hybrid virus-HE in Bm5 cells was significantly lower than that expressed in Sf21 cells. The titer of Ac-Bm hybrid virus-HE determined by plaque assays in Bm5 cells was similar to that determined in Sf21 cells, but the plaque size formed by Ac-Bm hybrid virus-HE in Bm5 cells was apparently smaller than that formed in Sf21 cells. In addition, expression levels and virus titers of Ac-Bm hybrid virus-LE in Sf21 and Bm5 were significantly lower than those of Ac-Bm hybrid virus-HE. Therefore, DNA sequences were determined for the region of the p143 gene controlling the host range in Ac-Bm hybrid viruses. The results showed that the deduced amino acid sequences of Ac-Bm hybrid virus-HE were almost identical to those of BmNPV. There were differences only in amino acids at positions 461 and 470, whereas those of Ac-Bm hybrid virus-LE were different at position 461, 470, 514, and 528 from those of BmNPV. In conclusion, our results clearly demonstrated that Ac-Bm hybrid virus-HE has an additional advantage of expanded host range for producing recombinant proteins.

Characterization of novel non-toxic Bacillus thuringiensis isolates from Korea.

Four Bacillus Thuringiensis isolates from soil samples produced parasporal inclusions which were non-toxic to insects. The isolates were named B. thuringiensis NTB-1, NTB-2, NTB-3 and NTB-4. The parasporal inclusions were shown to be ovoid by phase contrast and scanning electron microscopy. The serotypes of the four isolates were determined by agglutination using 33 antisera; NTB-1 and NTB-4 seemed to be subsp. israelensis, and NTB-2 seemed to be subsp. pondicheriensis. NTB-3 did not react with the 33 antisera. However, comparison of parasporal protein and plasmid DNA patterns of the four isolates with those of 15 known non-toxic B. thuringiensis strains demonstrated that the four isolates are novel.

A humanized antibody with specificity for hepatitis B surface antigen.

A murine monoclonal antibody H67 was characterized for the binding specificity, which showed that H67 recognizes a disulfide-bond-dependent conformational epitope of common a antigenic determinant on the hepatitis B surface antigen. The result suggested that this antibody may have the potential of replacing hepatitis B immune globulin in the prevention of hepatitis B virus (HBV) infection. Therefore, we have constructed the humanized antibody HuS10 by grafting the complementarity determining regions and some framework amino acid residues of H67 onto the most homologous human antibody variable regions, 21/28 for heavy chain variable region and B1 and J kappa 2 for light chain variable region, followed by combining with human constant regions C gamma 1 and C kappa. The affinity of the HuS10 was the same as that of the H67, 8 x 10(8) x 10(8)M-1, and the HuS10 neutralized the in vitro infection of adult human hepatocyte primary culture by adr or ayw subtype of HBV. The neutralization assay showed that the HuS10 had approximately 2,000-times higher specific activity than commercially available polyclonal HBIG. These results suggest that the humanized antibody will be useful in the prevention or treatment of HBV infection.

Characterization of a murine-human chimeric antibody with specificity for the pre-S2 surface antigen of hepatitis B virus expressed in baculovirus-infected insect cells.

We have produced and characterized a murine-human chimeric antibody with specificity for the pre-S2 surface antigen of hepatitis B virus (HBV) in baculovirus-infected insect cells. Recombinant baculovirus carrying the cDNA coding for the heavy or light chain of the chimeric antibody was constructed and co-infected into insect cells. The chimeric antibody (BV-S2) expressed in the cells was purified by an affinity chromatography on Protein A-Sepharose 4B column and characterized by N-terminal amino acid sequencing, affinity determination for pre-S2 peptide, endoglycosidase digestion and Clq binding assay, which were then compared with those of the chimeric antibody H69K that has the same amino acid sequence as BV-S2, but produced from transfected murine myeloma cells. The N-linked glycosylation of the BV-S2 antibody was also analyzed by culturing the baculovirus-infected cells in the presence of tunicamycin. The results showed that the BV-S2 was secreted following correct removal of the leader peptides, contained N-linked carbohydrate at the heavy chain, and had the same binding affinity and Clq binding ability as H69K, suggesting that the BV-S2 chimeric antibody is functional and thus may be useful in the prevention of HBV infection.

Expression and synergistic effect of three types of crystal protein genes in Bacillus thuringiensis.

Bacillus thuringiensis NT0423 newly isolated from sericultural farms in Korea produces quite atypical bipyramidal crystals of a common major band of ca. 130 kDa, and has dual specificity against Lepidoptera and Diptera. To enforce the Diptera toxicity of B. thuringiensis NT0423, cryIVD and cytA genes were transformed into B. thuringiensis NT0423. The transformant B. thuringiensis PT0529 was obtained from introduction of pCG5 into B. thuringiensis NT0423 by electroporation. The expression of crystal proteins in B. thuringiensis PT0529 was characterized by SDS-PAGE and electron microscopy. The results showed that crystalline inclusions of host, CryIVD and CytA were clearly observed in B. thuringiensis PT0529. Furthermore, the toxicity of B. thuringiensis PT0529 against Diptera was highly enforced by synergistic effect.

High level expression of hepatitis B virus preS1 peptide in Escherichia coli.

PreS1 region gene fragment encoding the N-terminal 56 amino acid (aa) of hepatitis B virus (HBV, adr subtype), which encodes B- and T-cell epitopes and an hepatocyte receptor binding site, was synthesized by PCR and fused to the 3'-end of MalE gene encoding maltose-binding protein (MBP) to yield expression plasmid pMalpreS1-56. The plasmid was introduced into Escherichia coli DH5 alpha and expressed at 37 degrees C under the control of inducible tac promoter. The resulting fusion protein was highly expressed in a soluble form, about 40% of total cellular proteins, but it bound only partially to an amylose column. Therefore, the soluble preS1 fusion protein was purified to near homogeneity by two passages of anion-exchange chromatography followed by gel filtration. The yield of the fusion protein was 70 mg per 1 culture that had been induced by IPTG for 6 h. The purified fusion protein was specifically cleaved by a Factor Xa digestion to release the preS1 peptide, which was then purified by gel filtration to homogeneity. The purity, integrity, antigenicity and immunogenicity of the purified preS1 peptide was confirmed by glycerol-SDS-PAGE, Western analysis, N-terminal amino acid sequencing and an indirect ELISA.

Cloning and characterization of cDNAs coding for heavy and light chains of a monoclonal antibody specific for the S antigen of hepatitis B virus.

The nucleotide (nt) sequences encoding the variable regions of the heavy and light chains of a murine monoclonal antibody (mAb) have been determined. The mAb recognizes a disulfide-bond-dependent conformational epitope on the hepatitis B virus surface antigen. The sequence analyses revealed that the variable regions of the heavy and light chains are members of mouse heavy-chain subgroup II(A) and kappa light-chain subgroup III, respectively.