Effect of insulin-induced hypoglycemia on the serum concentrations of thyroxine,triiodothyronine and reverse triiodothyronine

The effect of insulin-induced hypoglycemia on serum thyroid hormone concentrations was studied in nine healthy individuals. Before, during and after the hypoglycemia blood samples were taken for measurement of the concentrations of glucose, thyroxine (T₄), triiodothyronine (T₃), reverse triiodothyronine (rT₃), catecholamines and pituitary hormones.There was no change in the mean serum T₄ level (± the standard error of the mean) of 67 ± 2 μg/l. However, the T₃ concentrations rose from a mean basal level of 1.86 ± 0.06 μg/l to a mean peak of 2.51 ± 0.21 μg/l (P < 0.01) at 45 minutes after the insulin injection, and the rT₃ concentrations fell from a mean basal level of 0.184 ± 0.008 μg/l to a mean nadir of 0.171 ± 0.022 μg/l (not a significant change). The mean peak epinephrine level was 545 ± 103 ng/l and it occurred between 30 and 45 minutes after the insulin injection; the mean peak norepinephrine level was 584 ± 114 ng/l and it occurred between 30 and 90 minutes after the injection. The growth hormone levels reached a mean peak of 26.1 ± 4.8 μg/l and the plasma cortisol levels rose to 215 ± 9 μg/l. The mean basal prolactin level was 8.5 ± 0.9 μg/l; in five subjects there was a rise to a mean peak of 50.6 ± 14.6 μg/l, whereas in the remaining four no significant increase occurred. No correlation was found between the changes in the serum T₃ concentration and any of the other factors studied.It was concluded that acute hypoglycemia is associated with a rapid increase in the serum T₃ concentration.     

Plasma TSH and Serum T-4 Levels in Long-term Follow-up of Patients Treated with 131I for Thyrotoxicosis

In February 1972 58% of patients euthyroid after iodine-131 therapy given for thyrotoxicosis between 1954 and 1966 had a high plasma TSH (>7·4 μU/ml) and 42% a normal plasma TSH level. A group of 69 of the euthyroid patients with high plasma TSH levels (25·0±2·0 μU/ml) in 1972 were re-examined 15 and 24 months later. The mean plasma TSH in the 66 patients remaining euthyroid at 15 months was 22·6±1·8 μU/ml, while three patients had become hypothyroid. At 24 months 64 of the patients were still available for study, of whom 61 remained euthyroid with a mean plasma TSH of 21·6±2·0 μU/ml, and a further three had become hypothyroid.All of a group of 61 of the euthyroid patients with normal plasma TSH levels (4·0±0·2 μU/ml) in 1972 remained euthyroid at 24 months with a mean plasma TSH of 4·1±0·3 μU/ml, though the plasma TSH level had become slightly raised in three.The mean serum T-4 level in the euthyroid patients with a high plasma TSH was significantly lower, though still in the normal range, than that in the euthyroid patients with a normal plasma TSH both in 1972 and in 1974.Since no patient with a normal plasma TSH level after iodine-131 treatment six to 18 years earlier for thyrotoxicosis developed hypothyroidism over a two-year period, the follow-up of such patients need not be so rigorous as that of similarly treated euthyroid patients with raised plasma TSH levels in whom hypothyroidism developed at the rate of 5% per year.     

Blood Muramidase Activity in Colorectal Cancer

The serum muramidase levels were measured in 128 patients with primary or metastatic colorectal cancer, 166 tumour-free patients after resection of a colorectal cancer, and 172 controls. Muramidase levels over 10 μg/ml were detected in 30%-39% of the tumour-bearing patients, in 8·2% of the tumour free, and in only 1·7% of the controls (normal level 6·68 ± 1·42 μg/ml). Long-term follow up indicated that raised levels may occur as a transient phenomenon in recurrent or metastatic disease. The likely relation of abnormal serum muramidase activity and stimulation of the reticuloendothelial system is discussed.     

Effect of Vegetarianism and Smoking on Vitamin B12, Thiocyanate,and Folate Levels in the Blood of Normal Subjects

Vitamin B₁₂, thiocyanate, and folate levels in the blood were estimated in 69 apparently normal subjects, of whom 26 were non-vegetarian non-smokers, 19 non-vegetarian smokers, 15 vegetarian non-smokers, and nine vegetarian smokers. The serum total (cyanide-extracted) B₁₂ level (value A) ranged from 105 to 728 pg/ml, with a mean of 292 pg/ml. The highest values were found in non-vegetarian non-smokers and the lowest in vegetarian smokers. There was no significant difference in value A between smokers as a group and non-smokers as a group. On the other hand, in vegetarians value A was very significantly lower than in non-vegetarians regardless of their smoking habits.It is suggested that A may represent both the protein-bound and free forms of vitamin B₁₂ in the blood, and B mainly the free B₁₂, which may be the physiologically active form. The plasma thiocyanate level varied from 1·0 to 15 μmol/100 ml, being, as expected, much higher in smokers (mean 8·20 μmol/100 ml) than in non-smokers (mean 2·02 μmol/100 ml). There was a rough correlation between falling B₁₂ levels and rising thiocyanate levels. The serum folate level ranged from 2·75 to 15·75 ng/ml, and was slightly but significantly higher in vegetarians (mean 6·60 ng/ml) than in non-vegetarians (mean 4·79 ng/ml), reflecting the greater content of folate in a vegetarian diet.     

Measurement of Digoxin in Plasma and its use in Diagnosis of Digoxin Intoxication

A method for measuring the plasma-digoxin concentration uses the measurement of its inhibitory effect on ⁸⁶Rb uptake by human red cells in vitro. Patients receiving digoxin in whom there was no clinical evidence of digoxin intoxication had plasma digoxin concentrations ranging from 0·8 to 4·5 mμg./ml. Patients presenting with convincing clinical evidence of digoxin intoxication had plasma digoxin concentrations ranging from 4 to greater than 8 mμg./ml. It is suggested that the plasma digoxin concentration may be used as an aid in the diagnosis of digoxin intoxication.     

Studies on the congenitally goitrous sheep. The iodinated compounds of serum, and circulating thyroid-stimulating hormone

1. A group of normal and congenitally goitrous Merino sheep were investigated to identify the metabolic defect present in the abnormal animals. 2. Protein-bound iodine concentrations of serum from goitrous animals (average 5·7μg./100ml.) were higher than normal (average 4·2μg./100ml.; P 0·001), but the hormonal iodine measured as butanol-extractable ¹³¹I was low in the serum of goitrous (average 40·3% of protein-bound ¹³¹I) compared with that of normal (84·2%; P 0·02) sheep. The non-hormonal iodine of the serum of goitrous sheep appeared to include iodotyrosines and iodinated protein. 3. Starch-gel-electrophoretic separations of sera from normal and goitrous sheep after ¹³¹I injection (100–500μc) showed no qualitative differences in the radioactivity of protein components. No significant differences in thyroxine-binding in vitro by serum proteins of normal and goitrous sheep were observed. 4. The clearance rates of ¹³¹I-labelled iodotyrosines (t_½ 1·2–2·9hr.) and iodothyronines (t_½ 33·5–47·4hr.) were similar in normal and goitrous sheep. 5. The concentration of circulating thyroid-stimulating hormone was significantly higher (P<0·01 in three sheep, P<0·05 in one sheep) in goitrous sheep. 6. The congenital goitre appears to be due to compensatory hypertrophy of the gland resulting from an inability to synthesize an adequate supply of thyroid hormone.     

Simple Method for Detection of Infection of Peritoneum during Dialysis

The lysozyme (muramidase) content of peritoneal fluid samples has been found to be an early indicator of the onset of infection in the course of peritoneal dialysis. A level of 10·0 μg/ml indicates peritoneal infection and one of 7·5 μg/ml is highly suspicious.     

Glucose metabolism in the superovulated rat ovary in vitro. Effects of luteinizing hormone and the role of glucose metabolism in steroidogenesis

1. Superovulated rat ovary slices from rats treated with 20μg. of luteininzing hormone/100g. body wt. 2hr. before death and from control animals have been incubated in vitro. Output of Δ⁴-3-oxo steroids (0·2μmole/g. wet wt./hr. in control tissue) was linear for 4hr., and was increased by approx. 70% in slices from luteinizing hormone-treated rats. Rate of oxygen consumption (90·0±4·6μmoles/g. wet wt./hr.) was linear for 3hr. and unaltered by luteinizing hormone treatment or addition of glucose (1mg./ml.) to the medium. 2. In slices from control animals, steady-state rate of glucose uptake was 78·0±2·9μg. atoms of carbon/g. wet wt./hr.; steady-state rates of lactate output, pyruvate output and incorporation of [U-¹⁴C]-glucose carbon atoms into carbon dioxide and total lipid extract were 60·7±0·9, 2·4±0·1, 18·0±1·1 and 0·7±0·1μg. atom of carbon/g. wet wt./hr. and accounted for 104·5±1·9% of the glucose uptake. In slices from luteinizing hormone-treated rats, glucose uptake and outputs of lactate, pyruvate and [¹⁴C]carbon dioxide were increased by approx. 25%, and 108·4±3·2% of the glucose uptake could be accounted for. 3. The total lipid extract was separated by thin-layer chromatography and saponification. Of the ¹⁴C incorporated into this fraction during incubation with [U-¹⁴C]glucose 97% was found in the fractions containing glyceride glycerol and less than 3% in the fractions containing sterols, steroids or fatty acids. Appreciable quantities of ¹⁴C were incorporated into these lipid fractions from [1-¹⁴C]acetate. 4. From a consideration of the tissue glycogen content, the specific activities of [¹⁴C]lactate and glucose 6-phosphate (C-1) derived from [1-¹⁴C]-, [6-¹⁴C]- and [U-¹⁴C]-glucose, and the ratio of [¹⁴C]carbon dioxide yields from [1-¹⁴C]glucose and [6-¹⁴C]glucose, it was concluded that there was no appreciable glycogenolysis or flow through the pentose phosphate cycle. 5. In ovary slices from both control and luteinizing hormone-treated animals, glucose in vitro raised the incorporation rate of ¹⁴C from [1-¹⁴C]acetate into sterols and steroids. Luteinizing hormone in vivo stimulated the incorporation rate in vitro but only in the presence of glucose. 6. In slices incubated in medium containing [³H]water, [¹⁴C]sorbitol and glucose (1mg./ml.), the total water space (865±7·1μl./g.) and the extracellular water space (581±22μl./g.) were unchanged by luteinizing hormone treatment in vivo but the glucose space was raised from 540±23·6μl./g. to 639±31·3μl./g. 7. Luteinizing hormone treatment was found to lower the tissue concentration of the hexose monophosphates and to increase the total activity of hexokinase, glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase and possibly of phosphofructokinase. 8. The kinetic properties of a partially purified preparation of phosphofructokinase were found to be qualitatively similar to those from other mammalian tissues. 9. The results are discussed with reference to both the role of glucose metabolism in steroidogenesis and the mechanism by which luteinizing hormone increases the rate of glucose uptake.     

Serum and Urinary Folate in Liver Disease

During the active phase of viral hepatitis urinary folate loss was found to be 8·0 to 48·3 (mean 31·1) μg./day, compared with a normal urinary folate excretion of 0·1 to 18·0 (mean 9·5) μg./day. In cirrhosis and cardiac failure with congestive hepatomegaly the corresponding values were 25·8 to 55·0 (mean 35·7) μg./day and 2·5 to 61·6 (mean 26·9) μg./day, respectively. Urinary folate loss may be a significant factor in the aetiology of folate deficiency of chronic liver disease, particularly when dietary intake is poor.After prolonged dialysis in Visking casing urinary folate was almost totally dialysable, but an appreciable fraction of serum folate was not, even after 72 hours. The dialysable (free) folate fraction of serum and urine disappeared maximally during the first six hours'' dialysis, and was virtually cleared after 24 hours'' dialysis; clearance curves in normal individuals and in liver disease were comparable. The non-dialysable serum folate fraction was of similar magnitude in all subjects studied, in spite of marked variation in total folate, and probably represented protein-bound folate.     

Stimulation of adenine phosphoribosyltransferase by adenosine triphosphate and other nucleoside triphosphates

1. Adenine phosphoribosyltransferase was protected from inactivation on heating at 55° by the presence of 5-phosphoribosyl pyrophosphate. ATP, adenine, AMP or GMP had no protective effect on the activity of this enzyme. The presence of either 5-phosphoribosyl pyrophosphate or ATP did not protect adenine phosphoribosyltransferase against the loss of ATP stimulation obtained by heating at 55°. 2. At pH5·3 and 6·0 adenine phosphoribosyltransferase was stimulated by a narrow range of ATP concentration (15–25μm). At pH6·5 and 7·0 maximum stimulation was obtained with 25–30μm-ATP, and at pH7·4, 8·2 and 8·85 maximum stimulation was obtained over a wide range of ATP concentrations (60–200μm). With extracts that had been heated for 30min. at 55° no stimulation was observed at either pH5·3 or 7·4 with ATP concentrations up to 100μm. 3. Short periods of heating at 55° (1, 2 or 5min.) increased the stimulation of adenine phosphoribosyltransferase obtained with various concentrations of ATP. 4. The addition of CTP, GTP, deoxy-GTP, deoxy-TTP or XTP to assay mixtures resulted in weak stimulation of adenine-phosphoribosyltransferase activity. 5. It is suggested that there are at least three different forms of adenine phosphoribosyltransferase, each with a different affinity for ATP.     

A KATP Channel-Dependent Pathway within α Cells Regulates Glucagon Release from Both Rodent and Human Islets of Langerhans

Glucagon, secreted from pancreatic islet α cells, stimulates gluconeogenesis and liver glycogen breakdown. The mechanism regulating glucagon release is debated, and variously attributed to neuronal control, paracrine control by neighbouring β cells, or to an intrinsic glucose sensing by the α cells themselves. We examined hormone secretion and Ca²⁺ responses of α and β cells within intact rodent and human islets. Glucose-dependent suppression of glucagon release persisted when paracrine GABA or Zn²⁺ signalling was blocked, but was reversed by low concentrations (1–20 μM) of the ATP-sensitive K⁺ (K_ATP) channel opener diazoxide, which had no effect on insulin release or β cell responses. This effect was prevented by the K_ATP channel blocker tolbutamide (100 μM). Higher diazoxide concentrations (≥30 μM) decreased glucagon and insulin secretion, and α- and β-cell Ca²⁺ responses, in parallel. In the absence of glucose, tolbutamide at low concentrations (<1 μM) stimulated glucagon secretion, whereas high concentrations (>10 μM) were inhibitory. In the presence of a maximally inhibitory concentration of tolbutamide (0.5 mM), glucose had no additional suppressive effect. Downstream of the K_ATP channel, inhibition of voltage-gated Na⁺ (TTX) and N-type Ca²⁺ channels (ω-conotoxin), but not L-type Ca²⁺ channels (nifedipine), prevented glucagon secretion. Both the N-type Ca²⁺ channels and α-cell exocytosis were inactivated at depolarised membrane potentials. Rodent and human glucagon secretion is regulated by an α-cell K_ATP channel-dependent mechanism. We propose that elevated glucose reduces electrical activity and exocytosis via depolarisation-induced inactivation of ion channels involved in action potential firing and secretion.     

Asprin‐loaded strontium‐containing α‐calcium sulphate hemihydrate/nano‐hydroxyapatite composite promotes regeneration of critical bone defects

Our laboratory originally synthesized strontium(Sr)‐containing α‐calcium sulphate hemihydrate/nano‐hydroxyapatite composite (Sr‐α‐CSH/n‐HA) and demonstrated its ability to repair critical bone defects. This study attempted to incorporate aspirin into it to produce a better bone graft material for critical bone defects. After 5% Sr‐α‐CSH was prepared by coprecipitation and hydrothermal methods, it was mixed with aspirin solution of different concentrations (50 μg/ml, 200 μg/ml, 800 μg/ml and 3200 μg/ml) at a fixed liquid‐solid ratio (0.54 v/w) to obtain aspirin‐loaded Sr‐α‐CSH/n‐HA composite. In vitro experiments were performed on the composite extracts. The tibial defects (3 mm*5 mm) in SD rat model were filled with the composite for 4 weeks and 12 weeks to evaluate its osteogenic capacity in vivo. Our results showed its capability of proliferation, migration and osteogenesis of BMSCs in vitro got improved. In vivo treatment with 800 μg/ml aspirin–loaded Sr‐α‐CSH/n‐HA composite led to significantly more new bone formation in the defects compared with Sr‐α‐CSH/n‐HA composite and significantly promoted the expression of osteogenic‐related genes and inhibited osteoclast activity. In general, our research suggests that aspirin‐loaded Sr‐α‐CSH/n‐HA composite may have a greater capacity of repairing tibial defects in SD rats than simple Sr‐α‐CSH/n‐HA composite.     

Casein Fermentate of Lactobacillus animalis DPC6134 Contains a Range of Novel Propeptide Angiotensin-Converting Enzyme Inhibitors

This work evaluated the angiotensin-converting-enzyme (ACE)-inhibitory activities of a bovine sodium caseinate fermentate generated using the proteolytic capabilities of the porcine small intestinal isolate Lactobacillus animalis DPC6134 (NCIMB deposit 41355). The crude 10-kDa L. animalis DPC6134 fermentate exhibited ACE-inhibitory activity of 85.51% (±15%) and had a 50% inhibitory concentration (IC₅₀) of 0.8 mg protein/ml compared to captopril, which had an IC₅₀ value of 0.005 mg/ml. Fractionation of the crude L. animalis DPC6134 fermentate by membrane filtration and reversed-phase high-performance liquid chromatography (HPLC) generated three bioactive fractions from a total of 72 fractions. Fractions 10, 19, and 43 displayed ACE-inhibitory activity percentages of 67.53 (±15), 83.71 (±19), and 42.36 (±11), respectively, where ACE inhibition was determined with 80 μl of the fractions with protein concentrations of 0.5 mg/ml. HPLC and mass spectrometry analysis identified 25 distinct peptide sequences derived from α-, β-, and κ-caseins. In silico predictions, based on the C-terminal tetrapeptide sequences, suggested that peptide NIPPLTQTPVVVPPFIQ, corresponding to β-casein f(73-89); peptide IGSENSEKTTMP, corresponding to α_s1-casein f(201212); peptide SQSKVLPVPQ, corresponding to β-casein f(166-175); peptide MPFPKYPVEP, corresponding to β-casein f(124133); and peptide EPVLGPVRGPFP, corresponding to β-casein f(210-221), contained ACE-inhibitory activities. These peptides were chosen for chemical synthesis to confirm the ACE-inhibitory activity of the fractions. Chemically synthesized peptides displayed IC₅₀ values in the range of 92 μM to 790 μM. Additionally, a simulated gastrointestinal digestion confirmed that the ACE-inhibitory 10-kDa L. animalis DPC6134 fermentation was resistant to a cocktail of digestive enzymes found in the gastrointestinal tract.     

Modeling Reduction of Uranium U(VI) under Variable Sulfate Concentrations by Sulfate-Reducing Bacteria

The kinetics for the reduction of sulfate alone and for concurrent uranium [U(VI)] and sulfate reduction, by mixed and pure cultures of sulfate-reducing bacteria (SRB) at 21 ± 3°C were studied. The mixed culture contained the SRB Desulfovibrio vulgaris along with a Clostridium sp. determined via 16S ribosomal DNA analysis. The pure culture was Desulfovibrio desulfuricans (ATCC 7757). A zero-order model best fit the data for the reduction of sulfate from 0.1 to 10 mM. A lag time occurred below cell concentrations of 0.1 mg (dry weight) of cells/ml. For the mixed culture, average values for the maximum specific reaction rate, V_max, ranged from 2.4 ± 0.2 μmol of sulfate/mg (dry weight) of SRB · h⁻¹) at 0.25 mM sulfate to 5.0 ± 1.1 μmol of sulfate/mg (dry weight) of SRB · h⁻¹ at 10 mM sulfate (average cell concentration, 0.52 mg [dry weight]/ml). For the pure culture, V_max was 1.6 ± 0.2 μmol of sulfate/mg (dry weight) of SRB · h⁻¹ at 1 mM sulfate (0.29 mg [dry weight] of cells/ml). When both electron acceptors were present, sulfate reduction remained zero order for both cultures, while uranium reduction was first order, with rate constants of 0.071 ± 0.003 mg (dry weight) of cells/ml · min⁻¹ for the mixed culture and 0.137 ± 0.016 mg (dry weight) of cells/ml · min⁻¹ (U₀ = 1 mM) for the D. desulfuricans culture. Both cultures exhibited a faster rate of uranium reduction in the presence of sulfate and no lag time until the onset of U reduction in contrast to U alone. This kinetics information can be used to design an SRB-dominated biotreatment scheme for the removal of U(VI) from an aqueous source.     

Incorporation of Phosphatase Inhibitor in Culture Prompts Growth Initiation of Isolated Non-Growing Oocytes

In vitro folliculogenesis of primordial and early preantral follicles is necessary for increment of reproductive efficiency in domestic animals, humans and endangered species. Recent study in phosphatase and tensin homolog (Pten) -knockout mice has revealed that this phosphatase acts as an inhibitory factor in follicle activation of primordial pool with the resultant inhibition of oocyte growth. To test in vitro effect of a phosphatase inhibitor on growth initiation of isolated non-growing oocytes in neonatal ovaries, we applied a specific inhibitor (bpV (HOpic)) for PTEN in culturing system. Non-growing oocytes isolated from the ovaries of newborn BDF1 (C57BL/6 × DBA/2) pups were divided to four culture groups. Five days after culture, the oocytes in 14 μmol/l bpV only, 14 μmol/l bpV plus 100 ng/ml Kit Ligand (KL), and 100 ng/ml KL groups showed significantly (P<0.05) growth (19.3±0.55, 25.8±0.53 and 21.6±0.29 μm, respectively) compared with that of the control (no additive) (16.9±0.53 μm). In addition, western blotting in those groups showed enhanced expression of phosphorylated Akt. In conclusion, we clearly demonstrate that isolated non-growing oocytes develop in phosphatase inhibitor, especially to PTEN, incorporated culturing system, and show first as we know that oocytes with zona Pellucidae can be obtained in vitro from isolated non-growing oocytes.     

Continuous Intravenous Infusion of Small Doses of Insulin in Treatment of Diabetic Ketoacidosis

Continuous intravenous infusion of small amounts of insulin has been used in the management of a series of patients with diabetic ketoacidosis. In 13 patients with a plasma glucose level on admission of 725 mg/100 ml (± 80 S.E. of mean) and an arterial pH of 7·07 ± 0·05 a mean loading dose of 6·5 ± 0·82 units of soluble insulin was administered intravenously, and thereafter a sustaining infusion of 6·5 ± 0·82 U/hr was continued until ketosis was corrected and the plasma glucose fell below 300 mg/100 ml. The total insulin dose needed to achieve this was 39·2 ± 6·6 units given over a 3 to 10-hour period. Plasma insulin was measured in patients who had not previously received insulin and the mean level at an infusion rate of 4 U/hr was 75·6 ± 8·0 μU/ml. Plasma glucose fell at a regular rate of 101 ± 11 mg/100 ml/hr, and ketosis improved in parallel. Plasma potassium was well maintained throughout treatment. This regimen of treatment was clinically effective and simple to follow.     

The sodium, potassium and chloride of cerebral tissues: maintenance, change on stimulation and subsequent recovery

1. Cerebral tissues were prepared for incubation by cutting them from the brain rapidly and in situ, and the calcium concentration in the incubating medium was altered from the customary 2·8mm to 0·75mm. This provided incubated cerebral cortex with fluid and ion content more closely resembling that of the brain in vivo than hitherto obtained. 2. From a systematic difference in size between inulin spaces of slices with one and those with two cut surfaces, it was estimated that cutting directly affected a layer 0·02mm. thick. On the basis of the volume of this layer, it was calculated that the portion of the tissue not affected by cutting had an inulin space of 258 μl./g. initial wt., and during the process of preparation and incubation had gained 30μequiv. of sodium and 17 μequiv. of chloride/g. and had lost 14 μequiv. of potassium/g. 3. Several aspects of the ion content of the incubated tissue were compatible with the observed membrane potential of −60mv between cellular and extracellular phases. 4. In response to electrical stimulation, sodium of the non-inulin space increased from 28 to 57 μequiv./g., potassium decreased from 68 to 48 μequiv./g. and chloride increased from 16 to 22 μequiv./g. in the non-inulin space. These changes were complete in about 6min., and thereafter the concentrations remained steady during continued stimulation. Initial rates of change were 460 μequiv./g./hr. for sodium and 480 μequiv./g./hr. for potassium. 5. After stimulation was stopped the ionic composition of the tissue returned completely to its pre-stimulation state within 10min. Initial rates for extrusion of sodium and gain of potassium were 160 and 230 μequiv./g./hr. respectively.     

Cell-free DNA in Human Follicular Microenvironment: New Prognostic Biomarker to Predict in vitro Fertilization Outcomes

Cell-free DNA (cfDNA) fragments, detected in blood and in other biological fluids, are released from apoptotic and/or necrotic cells. CfDNA is currently used as biomarker for the detection of many diseases such as some cancers and gynecological and obstetrics disorders. In this study, we investigated if cfDNA levels in follicular fluid (FF) samples from in vitro fertilization (IVF) patients, could be related to their ovarian reserve status, controlled ovarian stimulation (COS) protocols and IVF outcomes. Therefore, 117 FF samples were collected from women (n = 117) undergoing IVF/Intra-cytoplasmic sperm injection (ICSI) procedure and cfDNA concentration was quantified by ALU-quantitative PCR. We found that cfDNA level was significantly higher in FF samples from patients with ovarian reserve disorders (low functional ovarian reserve or polycystic ovary syndrome) than from patients with normal ovarian reserve (2.7 ± 2.7 ng/μl versus 1.7 ± 2.3 ng/μl, respectively, p = 0.03). Likewise, FF cfDNA levels were significant more elevated in women who received long ovarian stimulation (> 10 days) or high total dose of gonadotropins (≥ 3000 IU/l) than in women who received short stimulation duration (7–10 days) or total dose of gonadotropins < 3000 IU/l (2.4 ± 2.8 ng/μl versus 1.5 ± 1.9 ng/μl, p = 0.008; 2.2 ± 2.3 ng/μl versus 1.5 ± 2.1 ng/μl, p = 0.01, respectively). Finally, FF cfDNA level was an independent and significant predictive factor for pregnancy outcome (adjusted odds ratio = 0.69 [0.5; 0.96], p = 0.03). In multivariate analysis, the Receiving Operator Curve (ROC) analysis showed that the performance of FF cfDNA in predicting clinical pregnancy reached 0.73 [0.66–0.87] with 88% specificity and 60% sensitivity. CfDNA might constitute a promising biomarker of follicular micro-environment quality which could be used to predict IVF prognosis and to enhance female infertility management.     

Photosynthetic electron transport in isolated maize bundle sheath cells

Fragments of bundle sheath strands, free of mesophyll cells and showing a chlorophyll a/b ratio of 6.0 to 6.6 were prepared from Zea mays by a mechanical method. They were unable to photoreduce ferricyanide but were able to photoreduce the membrane-permeant 2,5-dimethylquinone at a rate of 250 to 420 microequivalents per hour per mg chlorophyll (μeq/hr · mg Chl) at 21 C. In the presence of the catalase inhibitor KCN, methylviologen catalyzed a Mehler reaction at a rate of 120 to 180 μeq/hr · mg Chl. This was increased to 200 to 350 μeq/hr · mg Chl when the uncoupler methylamine was added. The rate of endogenous pseudocyclic electron flow, detected as a Mehler reaction, was also considerable (100 to 150 μeq/hr · mg Chl with methylamine). Diaminodurene supported a high rate of photosystem I-mediated electron flow to methylviologen (400 to 750 μeq/hr · mg Chl).