HPLC-MS/MS method for determination of betamethasone in human plasma with application to a dichorionic twin pregnancy pharmacokinetic and placental transfers studies

Abstract Betamethasone (BET) is a synthetic glucocorticoid recommended for pregnant women at imminent risk of preterm birth before 34 weeks to reduce neonatal complications. There are different techniques to describe BET plasma quantification. However, none quantified the plasmatic concentration of BET in dichorionic (DC) twin pregnancies using LC-MS. Our objectives were to develop and validate a method for quantifying BET by LC-MS for pharmacokinetic (PK) and placental transfer studies in DC twin pregnancies. Blood samples were collected after intramuscular administration of a single BET dose containing 6 mg disodium phosphate + 6 mg acetate. BET was determined in plasma by liquid-liquid extraction. The method showed linearity in the range of 2-250 ng/mL, as well as precision and accuracy with a coefficient of variation and relative standard errors ≤ 15%. Additionally, the method presented selectivity and did not present matrix or carry-over effect. Stability tests also presented coefficient of variation and relative standard errors ≤ 15%. This is the first study which describe maternal and fetal plasma concentrations of BET in a DC twin pregnancy. The BET PK parameters were AUC0-∞, CL/F, Vd/F, Cmax, Tmax of 292.20 h*ng/mL, 39.08 L/h, 278.72 L, 25.55 ng/mL and 0.58 h, respectively. The placental transfer ratios of umbilical vein/maternal vein and intervillous space/maternal vein were 0.14 and 0.19 and 0.40 and 0.27 for both twins, respectively. However, a clinical study with more subjects is imperative to confirm this higher concentration of BET in the intervillous space

Betamethasone suppresses the inflammatory response in LPS-stimulated dental pulp cells through inhibition of NF-κB.

OBJECTIVE: This study aimed to investigate the anti-inflammatory effect of betamethasone on LPS-stimulated human dental pulp stem cells (DPSCs) and its associated mechanism. The osteo-/odontogenic differentiation and osteoclast effect of betamethasone on DPSCs and stem cells from human exfoliated deciduous teeth (SHED) were evaluated. DESIGN: The proliferative effect of betamethasone on DPSCs was analyzed using a cholecystokinin octapeptide assay. The anti-inflammatory effect of betamethasone was investigated using quantitative polymerase chain reaction (qPCR) and ELISA. The anti-inflammatory mechanism was explored using qPCR, Western blot, and immunofluorescence staining. The osteo-/odontogenic differentiation and osteoclast effect of betamethasone on DPSCs and SHED were detected by qPCR. RESULTS: 1 µg L-1 betamethasone was found to have the strongest effect on DPSCs proliferation. The expression of pro-inflammatory cytokines and mediators, as well as prostaglandin E2 (PGE2) were significantly decreased following treatment with betamethasone in LPS- stimulated DPSCs. They were also decreased in response to an NF-κB inhibitor, Bay 11-7082. Betamethasone and Bay 11-7082 significantly inhibited the expression of p-p65 and promoted the nuclear exclusion of p65. Gene expression associated with osteo-/odontogenic differentiation was significantly up-regulated in betamethasone and osteogenic media (OM) treated groups. The ratio of the receptor activator of nuclear factor kappa B ligand (RANKL) and osteoprotegerin (OPG) at the mRNA level was suppressed in DPSCs and elevated in SHED. CONCLUSIONS: Betamethasone has an anti-inflammatory effect on LPS- stimulated DPSCs through a blockade of NF-κB activation and exhibits an osteo-/odonto-inductive effect on DPSCs and SHED. Although betamethasone displays an osteoclast effect on SHED.

Neuroprotective effects vary across nonsteroidal antiinflammatory drugs in a mouse model of developing excitotoxic brain injury.

Glutamate excitotoxicity is among the main cellular mechanisms leading to perinatal insults in human newborns. We used intracerebral injection of the glutamatergic glutamate N-methyl-D-aspartate-receptor agonist ibotenate to produce excitotoxic lesions mimicking the acquired white matter lesions seen in human preterm infants. We evaluated whether nonsteroidal antiinflammatory drugs (NSAIDs) protected against glutamate excitotoxicity. Aspirin (0.01-100 microg/d), indomethacin (0.1-10 microg/d), paracetamol (10-100 microg/d), or NS-398 (12.5 microg/d) was given daily before ibotenate (P1 to P5) or after ibotenate (P5 to P9). Lesion size was measured on Cresyl Violet-stained brain sections collected on P10. None of the drugs tested alone or in combination increased lesion size. Pretreatment with low- or high-dose aspirin and post-treatment with paracetamol or NS-398 protected against white matter lesions, whereas cortical lesions were decreased by pretreatment with low- or high-dose aspirin or post-treatment with NS-398. The corticosteroid betamethasone (0.18 microg/d) was neuroprotective when given before or after ibotenate and this effect was reversed by concomitant aspirin therapy (10 microg/d). In conclusion, perinatal NSAID administration may have beneficial effects on brain injury if appropriately timed.

Effect of transforming growth factor beta and basic fibroblast growth factor on steroid-impaired healing intestinal wounds.

A longitudinal intestinal wound model in the pig was used to assess the effect of parenteral steroids (betamethasone 12 mg 50 kg-1 intramuscularly twice daily) on breaking load. Steroid treatment significantly decreased the breaking load of wounds in the ileum and colon in comparison with wounds from saline-treated animals. In a further group of animals receiving steroids, paired longitudinal wounds were constructed. One wound of a pair was treated with a local application of transforming growth factor beta (TGF-beta) (5 micrograms per wound) or basic fibroblast growth factor (5 micrograms per wound) in a collagen suspension. The other wound was treated with a collagen suspension alone. Ileal wounds treated with TGF-beta were significantly stronger than collagen-treated controls at 7 days. The steroid-induced impairment of breaking load in intestinal wounds is partially reversed by a local application of TGF-beta in a collagen suspension at the time of surgery.

Exacerbation of pemphigus by rifampicin.

Rifampicin therapy caused exacerbation of skin lesions, rising serum pemphigus antibody titres, and decreased serum betamethasone levels in a 59-year-old man with pemphigus vulgaris. Exacerbation of pemphigus was confined to the period of rifampicin therapy and seems to be best explained by the effect of rifampicin on the metabolism of betamethasone. Physicians prescribing rifampicin for the treatment of tuberculosis should be aware of its potential to produce such an important adverse reaction.

RU 486 inhibits peripheral effects of glucocorticoids in humans.

RU 486 [17 beta-hydroxy-11 beta-(4-dimethylaminophenyl)17 alpha-(prop-1-ynyl)estra-4,9-dien-3-one] is a synthetic steroid receptor antagonist. To evaluate the peripheral antiglucocorticoid action of this compound, we investigated its ability to antagonize cutaneous steroid-induced vasoconstriction. This phenomenon, produced by three different topical steroids in six normal men, was consistently and significantly attenuated or abolished by oral administration of 6 mg/kg RU 486. This demonstration of a peripheral action of RU 486 is important in relation to the potential therapeutic use of this well tolerated drug in states of hypercortisolism. It also indicates that the cutaneous vasoconstrictor effects of topical steroids are mediated by occupancy of glucocorticoid receptors.

The influence of dimetindene maleate on the endogenous hydrocortisone synthesis suppressing potency of betamethasone.

Plasma hydrocortisone (cortisol) levels in 7 healthy volunteers were measured following administration of 0.25 mg betamethasone or 0.25 mg betamethasone and 1 mg dimetindene maleate. The H1-receptor antagonist dimetindene maleate did not influence the hydrocortisone suppressing activity of betamethasone, which was proved by comparison with the published plasma hydrocortisone levels of 162 young healthy untreated volunteers.

Topical betamethasone 17-valerate is an anticorticosteroid in the rat. 1. Dermal atrophy.

In the albino rat, topical betamethasone 17-valerate acts as an anticorticosteroid. This steroid is inactive in a dermal atrophy assay over a dose range where betamethasone and hydrocortisone display atrophogenic activity. At appropriate concentrations betamethasone 17-valerate competitively inhibits the atrophogenic effects of both betamethasone and triamcinolone acetonide. Since betamethasone and betamethasone 17-valerate penetrate rat skin in vivo at essentially the same rate, it is concluded that the latter compound is relatively resistant to hydrolysis during penetration, and that it binds to rat corticosteroid receptor proteins in such a manner as to prevent expression of corticosteroid activity. Therefore, the rat cannot be used as a model species to predict activity in man for this compound.