Among a number of antigens characterized in M. leprae, an etiological agent of Leprosy, the 18 kDa antigen, is unique to M. leprae. We have previously determined a sequence specific element in the 18 kDa gene of M. leprae, which confers transcriptional repression. In this report, we have examined if the element could be applied to genes other than the 18 kDa gene of M. leprae. To identify the roles of the regulatory sequence in heterologous promoter, we have constructed pB3 vector series, which contains BCG hsp65 promoter and the M. leprae 18 kDa transcription repression responsive element in tandem using LacZ gene as a reporter gene. Cloning of hsp65 promoters of M. bovis BCG or M. smegmatis in front of LacZ gene resulted in normal beta- galactosidase activity as expected. However, when the sequence element was placed between the promoter and the LacZ gene, beta-galactosidase activity was reduced 10-fold less. Also we have examined with pB3(-) vector, that harbors the transcription repression responsive element in a reversed orientation, the beta-galactosidase activity was found to be similar to pB3(+) vector. Thus, these results further confirm that M. leprae 18 kDa transcription repression responsive element could regulate BCG hsp65 heterologous promoter and that the element could act as an operator for the transcription of mycobacteria.
beta-Galactosidase ; Clone Cells ; Cloning, Organism ; Galactosidases ; Genes, Reporter ; Lac Operon ; Leprosy ; Mycobacterium bovis* ; Mycobacterium leprae ; Repression, Psychology*

BACKGROUND: It is currently thought that tissue valve degeneration is related to an animal's immune response, which is mainly due to cell surface alpha-Gal epitopes. Cell surface alpha-Gal epitopes are known to be degraded by the enzyme called green coffee bean alpha-galactosidase. It is also well known that alpha-Gal epitopes are immunologically stained by Griffonia Simplicifolia isolectin type B4. We know that many commercially available tissue valves are made of aortic valves and pericardial tissue of pig. So, we investigated whether alpha-Gal epitopes of the aortic valve and pericardial tissue of a pig can be removed by green coffee bean alpha-galactosidase, and we did so by comparing immunologic staining of the tissues before and after the enzyme treatment. MATERIAL AND METHOD: After treating fresh porcine aortic valve and pericardial tissue with green coffee bean alpha-galactosidase at concentrations of 0.5 unit/mL, 1.0 unit/mL, 2.0 unit/mL, respectively, under the condition of pH 6.5, temperature 4degrees C and 24 hours of incubation, each sample was stained with Griffonia Simplicifolia isolectin type B4 immunofluorescent labeling. We then examined whether the alpha-Gal epitopes were reduced or abolished in each consecutive concentration of green coffee bean alpha-galactosidase by comparing the degree of the Griffonia Simplicifolia isolectin B4 staining in each sample. RESULT: In the pig aortic valve tissue, a 1.0 unit/mL concentration of green coffee bean alpha-galactosidase at pH 6.5, 4degrees C and reaction for 24 hours was enough for complete removal of alpha-Gal epitopes from the cell surface on the immunostaining with Griffonia Simplicifolia isolectin B4. On the other hand, more alpha-Gal epitopes were present in the pig pericardial tissue on Griffonia Simplicifolia isolectin B4 staining before the enzyme treatment, and 1.0 unit/mL of galactosidase was not sufficient for complete removal of alpha-Gal from the tissue. 2.0 units/mL of green coffee bean alpha-galactosidase was needed to completely remove the alpha-Gal epitopes from the pericardial tissue on immunostaining. CONCLUSION: The alpha-Gal epitopes of the pig's aortic valve and pericardial tissue were successfully stained with Griffonia Simplicifolia isolectin B4. We could remove nearly all the alpha-Gal epitopes using green coffee bean alpha-galactosidase at the concentration of 1.0 unit/mL in the aortic valve of pig, and 2.0 unit/mL was need to nearly completely remove all the alpha-Gal epitopes in the pericardial tissue of pig under the condition of pH 6.5, 4degrees C and 24 hours of reaction time. In the near future, removal of alpha-Gal epitopes in the pig's aortic valve and pericardial tissue will become a powerful tool for the improvement of the tissue valve durability. It needs to be determined if alpha-galactosidase treated pig tissue is immune to human anti-Gal antibody or anti-Gal monoclonal antibodies.
alpha-Galactosidase ; Aortic Valve ; Coffee ; Epitopes ; Galactosidases ; Griffonia ; Hand ; Humans ; Hydrogen-Ion Concentration ; Lectins ; Pericardium ; Plant Lectins ; Reaction Time ; Tissue Transplantation

Fabry's disease is a rare, inborn error, sex-linked disorder of glycosphingolipid metabolism with death occurring from myocardial or renal involvement at 4th or 5th decades. The primary metabolic defect lies in the deficient tissue activity of the enzyme alpha-galactosidase A which results in progressive accumulation of the specific neutral glycosphingolipids, cerebroside dihexoside(CDH) and cerebroside triihexoside(CTH), within the lysosomes of endothelial, perithelial and smooth muscle cells of the cardiovascular and renal systems predominantly. Clinical manifestations are sequelae of the anatomic and physiologic alterations produced by the progressive deposition of glycosphingolipid in the tissues. We report the first case of successful renal transplantation in a patient with Fabry's disease in Korea. The patient was a 33-year-old male. Fabry's disease was confirmed by measurement of serum alpha- galactosidase level and renal biopsy. Biopsy finding showed lamellar inclusion bodies on electron microscopy. Galactosidase activity was also markedly decreased. He has been well for 49 months.
Adult ; alpha-Galactosidase ; Biopsy ; Fabry Disease* ; Galactosidases ; Humans ; Inclusion Bodies ; Kidney Transplantation* ; Korea ; Lysosomes ; Male ; Metabolism ; Microscopy, Electron ; Myocytes, Smooth Muscle ; Neutral Glycosphingolipids ; Transplantation

Fabry's disease is a rare, inborn error, sex-linked disorder of glycosphingolipid metabolism with death occurring from myocardial or renal involvement at 4th or 5th decades. The primary metabolic defect lies in the deficient tissue activity of the enzyme alpha-galactosidase A which results in progressive accumulation of the specific neutral glycosphingolipids, cerebroside dihexoside(CDH) and cerebroside triihexoside(CTH), within the lysosomes of endothelial, perithelial and smooth muscle cells of the cardiovascular and renal systems predominantly. Clinical manifestations are sequelae of the anatomic and physiologic alterations produced by the progressive deposition of glycosphingolipid in the tissues. We report the first case of successful renal transplantation in a patient with Fabry's disease in Korea. The patient was a 33-year-old male. Fabry's disease was confirmed by measurement of serum alpha- galactosidase level and renal biopsy. Biopsy finding showed lamellar inclusion bodies on electron microscopy. Galactosidase activity was also markedly decreased. He has been well for 49 months.
Adult ; alpha-Galactosidase ; Biopsy ; Fabry Disease* ; Galactosidases ; Humans ; Inclusion Bodies ; Kidney Transplantation* ; Korea ; Lysosomes ; Male ; Metabolism ; Microscopy, Electron ; Myocytes, Smooth Muscle ; Neutral Glycosphingolipids ; Transplantation

Galactosidase is generated from Aspergillus oryzae, which is widely used for antidiarrhea medicine to infants. Antibiotics and digestives were reported as a causative allergen inducing occupational asthma. Galatosidase-induced occupational asthma has not been reported yet. A forty-year-old female has suffered from rhinorrhea, sneezing, and nasal obstruction 1 year after handling galactosidase at obstetric and pediatric hospital, and then dyspnea appeared later. Skin prick test with inhalent allergens, beta-galactosidase, and Aspergillus oryzae showed strong positive reaction to beta-galactosidase only. Immunoinhibition test with beta-galactosidase and A. oryzae revealed inhibition to beta-galactosidase only. Bronchial provocation test with beta-galactosidase showed the dual asthmatic response. With these results, we confirmed that the patient has beta-galactosidase-induced occupational asthma and rhinitis.
Allergens ; Anti-Bacterial Agents ; Aspergillus oryzae ; Asthma, Occupational* ; beta-Galactosidase ; Bronchial Provocation Tests ; Dyspnea ; Female ; Galactosidases ; Hospitals, Pediatric ; Humans ; Infant ; Nasal Obstruction ; Oryza ; Rhinitis ; Skin ; Sneezing

A 21-year-old male suffered from generalized anhidrosis and heat intolerance for about 15 years. Other clinical features included multiple angiokeratoma on the back, scrotum and extremities, together with ocular involvement. The electronmicroscopic studies of the skin demonstrated laminated intracytoplamic inclusions in the endothelial cells, pericytes of dermal vessels, fibroblasts and eccrine glandular cells. a Galactosidase assay of leukocytes from the patient and his uncle showed nearly absence of the activity,
Angiokeratoma ; Endothelial Cells ; Extremities ; Fibroblasts ; Galactosidases ; Hot Temperature ; Humans ; Hypohidrosis* ; Leukocytes ; Male ; Pericytes ; Scrotum ; Skin ; Young Adult

OBJECTIVETo study the mechanism of lactose intolerance (LI) by cloning the mouse lactase cDNA and recombining a vector. METHODS Total murine RNA was isolated from the small intestine of a 4-week-old BALB/c mouse (d). Gene-specific primers were designed and synthesized according to the cDNA sequences of lactase-phlorizin hydrolase (LPH) in human, rat, and rabbit. A coding sequence (CDS) fragment was obtained using RT-PCR, and inserted into a clone vector pNEB-193, then the cDNA was sequenced and analyzed using bioinformatics.

RESULTSThe cDNA from the BALB/c mouse with 912 bp encoding 303 amino acid residues. Analysis of the deduced amino acid sequence using bioinformatics revealed that this cDNA shared extensive sequence homology with human LPH containing a conserved glycosyl hydrolase family 1 motif important for regulating lactase intolerance.

CONCLUSIONBALB/c mouse LPH cDNA (GenBank accession No: AY751548) provides a necessary foundation for study of the biological function and regulatory mechanism of the lactose intolerance in mice.

We report a new α-Galactosidase A (αGal-A) mutation in a 39-year-old Korean born, male Fabry disease patient. Fabry disease is a devastating, progressive inborn error of metabolism caused by X-linked genetic mutations. In this case, the first clinical symptom to occur was in childhood consisting of a burning pain originating in the extremities then radiating inwards to the limbs. This patient also stated to have ringing in his ears, angiokeratomas on his trunk, and cornea verticillata. He visited an outpatient cardiologist due to intermittent and atypical chest discomfort at the age of 39. Electrocardiographic and echocardiographic examination showed left ventricular hypertrophy. A physical examination revealed proteinuria without hematuria. The patient's plasma αGal-A activity was markedly lower than the mean value of the controls. After genetic counseling and obtaining written informed consent, we identified one hemizygous mutation in exon 4 of galactosidase alpha, c.617T>C (p.Leu206 Pro). He was eventually diagnosed as having Fabry disease.
Adult ; Angiokeratoma ; Burns ; Cornea ; Ear ; Echocardiography ; Electrocardiography ; Exons ; Extremities ; Fabry Disease* ; Galactosidases ; Genetic Counseling ; Hematuria ; Humans ; Hypertrophy, Left Ventricular ; Informed Consent ; Male ; Metabolism ; Outpatients ; Physical Examination ; Plasma ; Proteinuria ; Thorax

PURPOSE: To diagnose infantile colic from parent questionnaires, as well as investigating the risk factors and clinical course of infantile colic. METHODS: We retrospectively reviewed the medical records of 462 infants, with a corrected age of < 4 months at the time of visiting Inha University Hospital from January to December 2017. Parents responded to a 10-line questionnaire consisting of seven items relating to colic symptoms and three further items relating to underlying disease. The score was based on the number of days each symptom was evident during the preceding week. We defined infantile colic as the sum total being greater than seven points; if at least one of the three symptoms suggesting underlying disease was present, the infant was excluded from the diagnosis. RESULTS: One hundred and sixty-seven infants (36.1%) satisfied the criteria. The lower the gestational age, the more infantile colic they developed (P < 0.001). The prevalence of colic was higher in infants born with a birth weight < 2.5 kg (62.7% vs. 24.4%, P < 0.001) and in infants small for their gestational age, in the < 10th percentile (54.5% vs. 33.7%, P=0.003). The prevalence of colic was significantly different according to the type of feeding (P=0.001), being the lowest in breast-only feeding (29.8%), 32.8% in mixed feeding with breast milk and formula, and 49.7% in formula-only feeding. Colic symptoms improved by administering hydrolyzed formula (87.5%), low-lactose formula (47.1%), galactosidase (44.4%), and the probiotic Lactobacillus reuteri (34.5%). CONCLUSION: The prevalence of infantile colic was over 30%. Prematurity, lower birth weight, and small for gestational age were the risk factors of infantile colic. Clinical improvement was observed when active intervention was performed.
Birth Weight ; Colic ; Diagnosis ; Galactosidases ; Gestational Age ; Humans ; Infant ; Infant, Low Birth Weight ; Infant, Newborn ; Infant, Premature ; Lactobacillus reuteri ; Medical Records ; Milk, Human ; Parents ; Prevalence ; Probiotics ; Retrospective Studies ; Risk Factors

OBJECTIVE: The specific activity of lactase-phlorizin hydrolase (LPH) is very high at birth and sharply declines after weaning, producing lactose intolerance. The prevalence of lactose intolerance is up to 85% in Korean adults. Molecular basis of the regulatory mechanisms responsible for the decline of LPH specific activity is still unknown. In order to elucidate the molecular mechanisms regulating the LPH expression during development, LPH specific activity and mI4NA level of Korean fetal and adult intestines were compared. METHODS: 20 fetal small intestines (16-27 weeks) were obtained during therapeutic abortion and were divided into 3 equal length. 20 adult jejunal tissues were obtained from patients without small intestinal disease during laparotomy. Mucosal homogenates were prepared for dissacharidases specific activities measurement and total RNA was extracted for northern and slot hvbridization. LPH mRNA level was measured by laser densitometer. RESULTS: LPH specific activities of proximal, middle and distal portion of fetal intestines (n=20) were 36.2 +/- 22.5, 38.6 +/- 23.2 and 23.2 +/- 19.9 mu/mg protein, respectively. LPH specific activity of adult jejunum (n=8) was 5.9 +/- 1.8 mu/mg protein and significantly (p<0.05) lower than those of fetal intestines. However, there was no significant difference in sucrase and trehalase specific activities between fetal intestines and adult jejunum. Although LPH specific activity of adult jejunum was lower than those of fetal intestines, LPH mBNA level of adult jejunum was as high as those of fetal intestines. CONCLUSION: These results show that LPH specific activity and mRNA level do not parallel, indicating the posttranscriptional control of fetal development of LPH expression.
Abortion, Therapeutic ; Adult* ; Female ; Fetal Development ; Fetus* ; Humans ; Intestinal Diseases ; Intestine, Small* ; Intestines ; Jejunum ; Lactase* ; Lactase-Phlorizin Hydrolase ; Lactose Intolerance ; Laparotomy ; Parturition ; Pregnancy ; Prevalence ; RNA ; RNA, Messenger* ; Sucrase ; Trehalase ; Weaning