Enhanced biomass production of duckweeds by inoculating a plant growth-promoting bacterium, Acinetobacter calcoaceticus P23, in sterile medium and non-sterile environmental waters.

Duckweed offers the promise of a co-benefit culture combining water purification with biomass production. Acinetobacter calcoaceticus P23 is a plant growth-promoting bacterium isolated from a duckweed, Lemna aequinoctialis. This study quantified its growth-promoting effect on three duckweeds (L. aoukikusa, L. minor, and Spirodela polyrhiza) in sterile Hoagland solution and evaluated its usefulness in duckweed culture under non-sterile conditions. P23 promoted growth of three duckweeds in sterile Hoagland solution at low to high nutrient concentrations (1.25-10 mg NO3-N/L and 0.25-2.0 mg PO4-P/L). It increased the biomass production of L. aequinoctialis 3.8-4.3-fold, of L. minor 2.3-3.3-fold, and of S. polyrhiza 1.4-1.5-fold after 7 days compared with noninoculated controls. P23 also increased the biomass production of L. minor 2.4-fold in pond water and 1.7-fold in secondary effluent of a sewage treatment plant under non-sterile conditions at laboratory-scale experiments. P23 rescued L. minor from growth inhibition caused by microorganisms indigenous to the pond water. The results demonstrate that the use of P23 in duckweed culture can improve the efficiency of duckweed biomass production, and a positive effect of P23 on duckweed-based wastewater treatment can be assumed.

Different X-linked KDM5C mutations in affected male siblings: is maternal reversion error involved?

Genetic reversion is the phenomenon of spontaneous gene correction by which gene function is partially or completely rescued. However, it is unknown whether this mechanism always correctly repairs mutations, or is prone to error. We investigated a family of three boys with intellectual disability, and among them we identified two different mutations in KDM5C, located at Xp11.22, using whole-exome sequencing. Two affected boys have c.633delG and the other has c.631delC. We also confirmed de novo germline (c.631delC) and low-prevalence somatic (c.633delG) mutations in their mother. The two mutations are present on the same maternal haplotype, suggesting that a postzygotic somatic mutation or a reversion error occurred at an early embryonic stage in the mother, leading to switched KDM5C mutations in the affected siblings. This event is extremely unlikely to arise spontaneously (with an estimated probability of 0.39-7.5 × 10(-28) ), thus a possible reversion error is proposed here to explain this event. This study provides evidence for reversion error as a novel mechanism for the generation of somatic mutations in human diseases.

Age related expression of Werner's syndrome protein in selected tissues and coexpression of transcription factors.

AIMS: Werner's syndrome (WS) is an uncommon autosomal recessive disease resulting from mutational inactivation of human WRN helicase, Werner's syndrome protein (WRNp). Patients with WS progressively develop a variety of aging characteristics after puberty. The aim of this study was to determine the distribution of WRNp and the expression of the transcription factors regulating WRN gene expression in a variety of human organs in an attempt to understand the WS phenotype. METHODS: Tissue specimens were obtained from 16 controls aged from 27 gestational weeks to 70 years of age and a 56 year old female patient with WS. The distribution of WRNp and the expression of the transcription factors regulating WRN gene expression-SP1, AP2, and retinoblastoma protein (Rb)- were studied in the various human organs by immunohistochemical and immunoblot analyses. RESULTS: In the healthy controls after puberty, high expression of WRNp was detected in seminiferous epithelial cells and Leydig cells in the testis, glandular acini in the pancreas, and the zona fasciculata and zona reticularis in the adrenal cortex. In addition, the SP1 and AP2 transcription factors, which regulate WRNp gene expression, appeared in an age dependent manner in those regions where WRNp was expressed. In controls after puberty, SP1 was expressed in the testis and adrenal gland, whereas AP2 was expressed in the pancreas. CONCLUSIONS: These findings suggest that the age specific onset of WS may be related to age dependent expression of WRNp in specific organs.

Development of GABAergic neurons and their transporter in human temporal cortex.

The gamma-aminobutyric acid (GABA) system plays an important role in the early development of the hippocampal formation. The immunohistochemical expression of gamma-aminobutyric acid transporter-1, GAT-1, in the human developing temporal cortex was examined, and the distribution of GAT-1 was compared with that of the 67-kDa isoform of glutamic acid decarboxylase as a marker of GABAergic neurons. Four postmortem tissue specimens from young patients with hippocampal sclerosis were also examined. GAT-1 immunoreactivity was present, with a few puncta, in the neuropil of the stratum oriens and in the molecular layer of the dentate gyrus from 21-22 weeks of gestation, and in the stratum lacunosum-moleculare from 26 weeks of gestation. The peak expression of GAT-1 was seen in early infancy and that of glutamic acid decarboxylase in the perinatal period. These findings may reflect the development of GABAergic inhibitory systems, and may be related to the seizure susceptibility in infancy and early childhood. In the temporal lobes with hippocampal sclerosis, GAT-1 immunoreactivity of the neuropil was preserved in the vicinity of the neuronal loss of the hippocampus. This finding may result from the neurotrophic function of GAT-1 and may be related to its ability of neuronal repair and plasticity in childhood.

The effect of heat-treated human bone morphogenetic protein on clinical implantation.

For the clinical usage of human-derived bones, it is necessary to treat bones to reduce the risk of contamination by microorganisms. Bone morphogenetic protein is vulnerable to chemicals, but shows resistance to thermal heat to 70 degrees C in a short time. In this experiment, crude human bone morphogenetic protein was extracted from heat-treated bones at 60 degrees C for 10 hours and from nonheated bones. Sodium dodecyl sulfate polyacrylamide gel electrophoresis for these specimens was done. Gelatin capsules containing 5 mg of crude human bone morphogenetic protein extracted from heated and nonheated bones were implanted into thigh muscle pouches of five mice. At 20 days after implantation, the heterotopic bone formation was compared by evaluating the radiographic and histologic analyses. The sodium dodecyl sulfate polyacrylamide gel electrophoresis pattern of the human bone morphogenetic proteins showed five main bands (16, 22, 28, 35, and 67 kDa) that were almost identical. Heterotopic bone formation observed on the radiograph was induced by crude human bone morphogenetic protein from heated bones in a manner similar to that used for nonheated bones. The results from this study show that heat-treated bone preserves osteoinduction.

Characteristic features of the acetabular labrum in healthy children.

Radial magnetic resonance images of the acetabular labrum were obtained on 40 hips of healthy children. There were no right-left or male-female differences. In children aged 11 years or younger, the labrum on the antero-superior weightbearing portion was triangular in shape, and there was an insular-shaped or linear high-intensity area inside; on the mid-superior portion, the labrum appeared as a regular triangular, low-intensity area; and on the postero-superior portion, it was flat. In children aged 12 and 13 years, the shape of the labrum in each portion was similar to that of the younger children, but the high signal intensity area on the antero-superior portion appeared less frequently. The size of the labrum relative to the femoral head was greater in younger children.

Postoperative magnetic resonance imaging of lumbar disc herniation: comparison of microendoscopic discectomy and Love's method.

STUDY DESIGN: We performed a study to compare the magnetic resonance imaging findings up to 24 weeks after microendoscopic discectomy or surgery using Love's method in patients with lumbar disc herniation. OBJECTIVES: The objective was to determine whether or not microendoscopic discectomy was minimally invasive with respect to the nerve roots, cauda equina, and paravertebral muscles by comparing the postoperative magnetic resonance imaging findings in patients treated by microendoscopic discectomy and the conventional Love's method. SUMMARY OF BACKGROUND DATA: We introduced microendoscopic discectomy as a minimally invasive surgical procedure for lumbar disc herniation in September 1998 and have obtained good results. Microendoscopic discectomy is superior to the conventional Love's method in that it reduces postoperative pain, shortens the duration of hospitalization, and allows earlier resumption of normal activities. However, the effect of microendoscopic discectomy on the nerves and paravertebral muscles has not been evaluated objectively. METHODS: Enhancement of the nerve roots and paravertebral muscles, as well as the configuration of the cauda equina at the level of herniation, was assessed on axial magnetic resonance images obtained with contrast enhancement using gadolinium-diethylenetriamine penta-acetic acid before surgery and 1, 4, 8, 12, and 24 weeks after surgery in 25 patients who underwent microendoscopic discectomy and 15 patients who were treated using Love's method. RESULTS: Increased enhancement of the nerve roots was seen in 50.0% of the microendoscopic discectomy group and 46.2% of the Love group at 1 week after surgery. Enhancement of the paravertebral muscles at the surgical site tended to persist for longer in the microendoscopic discectomy group than in the Love group. However, muscle enhancement was widespread in some patients from the Love group. Abnormalities of the cauda equina attributed to surgical invasion were seen in 12.5% of the microscopic discectomy group and 15.4% of the Love group at 1 week after surgery. CONCLUSIONS: Microendoscopic discectomy had an effect on the nerve roots and cauda equina that was comparable with that of Love's method. The magnetic resonance images of the route of entry failed to show that microendoscopic discectomy is appreciably less invasive with respect to the paravertebral muscles.

Immunohistochemical expression and pathogenesis of BLM in the human brain and visceral organs.

Bloom syndrome (BS) involves the clinical features of telangiectatic erythema, immunodeficiency, and an increased risk for cancer. In order to clarify the pathogenetic significance of the responsible gene, BLM, which encodes a protein possessing homology to Escherichia coli RecQ helicase, the immunohistochemistry of BLM was examined in human brains and visceral organs from fetuses to adults and an adult with BS, using anti-BLM antibodies. Purkinje cells exhibited positive BLM immunoreactivity from 21 gestational weeks (GW), which transiently increased at approximately 40 GW. Neurons of the pontine tegmentum were immunolabeled from the early fetal period. In visceral organs, positive BLM immunoreactivity was observed in the Hassal corpuscles in the thymus from 24 GW, in beta-cells in the Langerhans islets of the pancreas from 36 GW, and in sperm cells and sperms of the testes from 11 years of age. But in a patient with BS, it was negative in the pancreas and testis tissues examined. The characteristic effect of BLM on specific cells in different periods suggests that the BLM gene product is closely related to neuronal development as well as immune, insulin secretory and sperm functions, which appear in different periods, and disorders of which are major symptoms of BS.

Two hyperandrogenic adolescent girls with congenital portosystemic shunt.

UNLABELLED: We describe two adolescent girls with a congenital portosystemic shunt who exhibited hyperandrogenism in addition to insulin resistant hyperinsulinaemia. Case 1 was referred to our clinic to undergo a routine clinical work-up prior to tonsillectomy at 14 years of age. Mild liver dysfunction was identified and hypogenesis of the portal vein with a congenital portosystemic shunt diagnosed. Primary amenorrhoea and virilization were evident and an endocrinological evaluation revealed hyperandrogenism and insulin resistant hyperinsulinaemia. Case 2 was referred at 15 years of age because of cardiomegaly. Mild liver dysfunction and hyperbilirubinaemia led to a diagnosis of agenesis of the portal vein with a congenital portosystemic shunt. Virilization was evident and an endocrinological evaluation revealed hyperandrogenism and insulin resistant hyperinsulinaemia. The haemodynamics of these patients were similar to those of secondary portosystemic shunt due to liver cirrhosis, which is often associated with hyperinsulinaemia and/or non-insulin dependent diabetes mellitus. On the other hand, hyperandrogenism is associated with certain insulin-resistant conditions with hyperinsulinaemia, including the polycystic ovary syndrome (PCO). Hyperinsulinaemia is believed to cause hyperandrogenism in patients with PCO by stimulating androgen production in both the ovary and adrenal gland. Therefore, in congenital portosystemic shunts, hyperinsulinaemia is also thought to cause hyperandrogenism due to the same mechanism. CONCLUSION: A certain percentage of female patients with hyperandrogenism, likely including those with polycystic ovary syndrome may also have congenital portosystemic shunts. Our results indicate that serum levels of total bile acids and ammonia are prognostic indicators of this hepatic vascular anomaly.

Significance of cathepsin B accumulation in synovial fluid of rheumatoid arthritis.

We measured and compared the activities of various kinds of proteinases, such as cysteine, serine, aspartic, and metalloproteinases, in synovial fluids of 16 patients with rheumatoid arthritis (RA) and 18 patients with osteoarthritis (OA). More than 19-fold higher activity of cathepsin B and about 6-fold higher activity of prolylendopeptidase, compared to those of OA, were accumulated in RA fluid. Moreover, levels of cathepsins B and S using the corresponding sandwich enzyme immunoassays were statistically higher in RA fluid than those in OA. Significant amounts of 41-kDa and 35-kDa procathepsin L were detected in RA fluid using gelatin zymography, while 41-kDa enzyme alone was detected in OA. Cathepsin B in RA fluid could degrade collagen, and this degradation was suppressed by the addition of CA-074, a specific inhibitor of cathepsin B. Therefore, cathepsin B may participate in joint destruction of RA, and its inhibitor may be effective for RA care.

Early immunohistochemical detection of axonal damage and glial activation in extremely immature brains with periventricular leukomalacia.

Extremely low birth weight (ELBW) infants, who died at 12 hours to 7 days after birth, with periventricular leukomalacia (PVL), were examined by means of neuropathological and immunohistochemical methods. Fourteen infants without PVL were used as controls. Anti-beta-amyloid precursor protein (APP), glial fibrillary acidic protein (GFAP), and ionized calcium-binding adaptor molecule 1 (Iba1) antibodies were used as markers for axonal damage, reactive astrocytes and activated microglia, respectively. Thirteen of 14 ELBW infants with PVL showed a widespread distribution of leukomalacia and 10 showed postnatal-onset of leukomalacia. In 12 of the 14 infants with PVL, regions of APP-reactive axons were found multifocally in the cerebral white matter, but 8 of them did not show coagulation necrosis on HE staining. GFAP-positive cells and Iba1-positive cells were markedly found in the white matter of all cases with PVL and slightly in all 14 controls. These results indicated that in ELBW infants, the distribution and formation of PVL foci are widespread and characteristic and so may involve motor and intellectual abilities in ELBW infants. Therefore, the perinatal management to maintain an appropriate cerebral circulation and oxygenation may be very important.

Radial-sequence magnetic resonance imaging in evaluation of acetabular labrum.

We investigated the usefulness of a radial-sequence magnetic resonance imaging (MRI) technique in the visualization of the acetabular labrum, which surrounds the acetabulum. In 22 hip joints of 12 volunteers, T2-weighted images were obtained on 24 radial planes of the acetabular rim, set at 15 degrees -intervals, using the small tip angle gradient echo method. We examined 7 planes in the weight-bearing portion. The acetabular labrum in the weight-bearing portion was depicted in good contrast to the surrounding tissues. The shape of the labrum differed among individuals and also in the anterior and posterior portions of the labrum. The signal intensity of the labrum was low or partially moderate. There was a high signal intensity band on the base of the acetabular labrum in several portions, which should be carefully interpreted to avoid confusion with abnormality. We concluded that radial-sequence MRI could be a useful technique for evaluation of the condition of the acetabular labrum in the weight-bearing portion.

[Consent to enrollment in randomized clinical trials].

Multi-facility joint randomized clinical trials for cancer treatment are the most common method of current clinical study. However, the difference in the rate of participation in multi-facility randomized clinical trials may damage the resulting general validity. Therefore, we studied whether the rate of participation in randomized clinical trials is different between university hospitals and other general hospitals using a questionnaire (with anonymity preserved). There were 744 subjects from university hospitals and 339 from general hospitals participating in the study. The results showed that 10.9 percent of those from university hospitals were willing to participate, against 28.1 percent who were not. Of those from general hospitals (public hospitals in this study) 10.6 percent answered that they would participate and 27.4 percent that they would not. Little difference was found in the rate of participation in randomized clinical trials between university hospitals and other general hospitals. The focus is thus on doctors to solve the possible differences between the facilities in further multi-facility joint studies.

MR imaging of perinatal brain damage: comparison of clinical outcome with initial and follow-up MR findings.

BACKGROUND AND PURPOSE: The purpose of our study was to determine whether MR studies in the neonatal period are predictive of the neuroradiologic sequelae and clinical outcome in premature and term infants with perinatal brain injury. METHODS: Thirty subjects (15 premature and 15 term infants) with abnormalities revealed by initial MR studies were reexamined approximately 1 year after birth with both MR imaging and a neurologic assessment. All initial MR studies were performed between 35 and 45 weeks corrected age in premature infants and within 28 days of life in term infants. The initial MR studies were evaluated for deep gray matter involvement, hemispheric parenchymal change, intracranial hemorrhage, and periventricular signal and/or morphologic changes. These MR findings were compared with the follow-up MR findings and with the neurologic outcome. RESULTS: The development of cerebral palsy in premature infants was related to the following initial MR findings: subependymal hemorrhage associated with parenchymal destruction, periventricular signal alteration with irregularity of the ventricular wall, and widespread cerebral infarction. These MR findings were predictive of the subtypes of cerebral palsy. In term asphyxiated infants, T2 signal alterations of the deep gray matter rather than T1 shortening and diffuse involvement of the hemispheres were predictive of an unfavorable outcome. Both in term and premature infants, focal hemispheric parenchymal lesions alone (including infarction and intracerebral, subdural, intraventricular, and subarachnoid hemorrhage) did not produce poor outcomes. CONCLUSION: MR studies performed at or near term in either premature or term infants with perinatal brain damage are effective in predicting both late neuroradiologic and clinical outcome.

Serum hyaluronic acid level reflects volume and ATP levels of the liver after extended hepatectomy with and without preoperative portal vein occlusion.

The purpose of this study was to examine the hypothesis that the serum hyaluronic acid (HA) level reflects the volume and adenosine triphosphate (ATP) levels of the liver after extended hepatectomy (ExHx) with and without preoperative portal vein occlusion (PVO). Rats were randomly divided into two groups, PVO-ExHx and sham-ExHx. At the first stage, they underwent PVO or sham surgery by an occlusion of the portal vein supplying the left lateral and median lobes or by similar manipulation but without PVO, respectively. Seven days after first-stage surgery, both groups received ExHx exceeding portal vein-occluded lobes, which was the excision of the left lateral, median, and right lateral lobes of the liver reported as a 90% hepatectomy model. On Days 0, 1, 2, and 3 after ExHx, the serum HA concentrations, liver weights, and hepatic ATP levels were determined. Liver volumes were restored to similar levels in both groups, but restoration was faster in the PVO-ExHx group because preoperative PVO allowed the excised liver less volume. Lower serum HA levels were significantly associated with significantly higher hepatic ATP levels and with a lower mortality rate in the PVO-ExHx group. The serum HA level correlated significantly (P < 0.001) with the liver weight and hepatic ATP level. The serum HA level serves as a proxy for clinically important parameters following major hepatic surgery.

Hepatic adenine nucleotides and microsomal cholesterol 7 alpha-hydroxylase activity in the obstructed and freely draining lobes of the liver after selective bile duct obstruction.

BACKGROUND: The effect of selective bile duct obstruction (SBDO) on hepatic reserve function of the bile duct obstructed (BDO) and nonobstructed freely draining (FD) lobes of the liver is obscure. METHODS: The bile duct branches draining from the left lateral and median lobes of the liver were ligated for 4 and 10 days in rats, and hepatic reserve functions in BDO and FD lobes were assessed by microsomal cholesterol 7 alpha-hydroxylase activities and by hepatic adenine nucleotide and energy charge levels. The values were compared with those in sham-operated control liver. Cholesterol 7 alpha-hydroxylase activities were determined by gas-liquid chromatography-mass spectrometry, and hepatic adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) levels with high-pressure liquid chromatography. RESULTS: The histological examination of the BDO lobes showed proliferation and formation of new bile ductules and fibrous connective tissue linking portal areas. Microsomal cholesterol 7 alpha-hydroxylase activities, hepatic energy charge and adenine nucleotide levels did not differ between FD and BDO lobes, and the values were similar to those in the sham-operated liver. CONCLUSIONS: Selective bile duct obstruction shows no adverse effects on microsomal and mitochondrial functions in either BDO or FD lobes of the liver.

Hepatic adenine nucleotides and microsomal cholesterol 7 alpha-hydroxylase activity in the obstructed and freely draining lobes of the liver after selective bile duct obstruction.

BACKGROUND: The effect of selective bile duct obstruction (SBDO) on hepatic reserve function of the bile duct obstructed (BDO) and nonobstructed freely draining (FD) lobes of the liver is obscure. METHODS: The bile duct branches draining from the left lateral and median lobes of the liver were ligated for 4 and 10 days in rats, and hepatic reserve functions in BDO and FD lobes were assessed by microsomal cholesterol 7 alpha-hydroxylase activities and by hepatic adenine nucleotides and energy charge levels. The values were compared with those in the sham-operated control liver. Cholesterol 7 alpha-hydroxylase activities were determined by gas-liquid chromatography--mass spectrometry, and hepatic adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) levels with high-pressure liquid chromatography. RESULTS: The histological examination of the BDO lobes showed proliferation and formation of new bile ductules and fibrous connective tissues linking portal areas. Microsomal cholesterol 7 alpha-hydroxylase activities, hepatic energy charge and each adenine nucleotide level did not differ between FD and BDO lobes, and the values were similar to those in the sham-operated liver. CONCLUSIONS: Selective bile duct obstruction shows no adverse effects on microsomal and mitochondrial functions in both the BDO and FD lobes of the liver.