A randomised controlled feasibility trial of a clinical protocol to manage hypotension during major non-cardiac surgery.

Intra-operative hypotension is a risk factor for postoperative morbidity and mortality. Minimally invasive monitors that derive other haemodynamic parameters, such as stroke volume, may better inform the management of hypotension. As a prelude to a multicentre randomised controlled trial, we conducted a single-centre feasibility trial of a protocol to treat hypotension as informed by minimally invasive haemodynamic monitoring during non-cardiac surgery. We recruited adults aged ≥40 years with cardiovascular risk factors who underwent non-cardiac surgery requiring invasive arterial pressure monitoring. Participants were randomly allocated to usual care, or a clinical protocol informed by an arterial waveform contour analysis monitor. Participants, outcome assessors, clinicians outside operating theatres and analysts were blinded to treatment allocation. Feasibility was evaluated based on: consent rate; recruitment rate; structured feedback from anaesthesia providers; and between-group differences in blood pressure, processes-of-care and outcomes. The consent rate among eligible patients was 33%, with 30 participants randomly allocated to the protocol and 30 to usual care. Anaesthesia providers rated the protocol to be feasible and acceptable. The protocol was associated with reduced fluid balance and hypotension exposure in the peri-operative setting. Postoperative complications included: acute myocardial injury in 18 (30%); acute kidney injury in 17 (28%); and surgical site infection in 7 (12%). The severity of complications was rated as moderate or severe in 25 (42%) participants. In summary, this single-centre study confirmed the feasibility of a multicentre trial to assess the efficacy and safety of a physiologically guided treatment protocol for intra-operative hypotension based on minimally invasive haemodynamic monitors.

Predictors of Post-operative Pain and Opioid Consumption in Patients Undergoing Liver Surgery.

BACKGROUND: Post-operative pain management is a critical component of perioperative care. Patients at risk of poorly controlled post-operative pain may benefit from early measures to optimize pain management. We sought to identify risk factors for post-operative pain and opioid consumption in patients undergoing liver resection. METHODS: This is a multi-institutional prospective nested cohort study of patients undergoing open liver resection. Opioid consumption and pain scores were collected following surgery. To estimate the effects of patient factors on opioid consumption (oral morphine equivalents-OME) and on pain scores (NRS-11), we used generalized linear models and multivariable linear regression model, respectively. RESULTS: One hundred and fifty-three patients who underwent open liver resection between 2013 and 2016 were included in the study. The mean patient age was 62.2 years, and 43.3% were female. Younger patients were significantly more likely to use more opioids in the early post-operative period (16.7 OME/10 years, p < 0.001). Patient factors that were significantly associated with increased NRS-11 pain scores also included younger patient age (difference in pain score of 0.3/10 years with cough and 0.2/10 years at rest, p < 0.01 for both) as well as a history of analgesic use (difference in pain score of 0.9 with cough and 0.6 at rest, p < 0.01 and p = 0.07, respectively). CONCLUSION: Younger patients and those with a history of analgesic use are more likely to report higher post-operative pain and require higher doses of opioids. Early identification of these patients, and measures to better manage their pain, may contribute to optimal perioperative care.

Use of prothrombin complex concentrate for management of coagulopathy after cardiac surgery: a propensity score matched comparison to plasma.

BACKGROUND: An important cause of coagulopathy in cardiac surgery is impaired thrombin generation. While plasma is often used to correct this element of the coagulopathy, studies in vitro suggest that prothrombin complex concentrates (PCCs) might be more effective. Comparative data, however, are scant. METHODS: We compared the outcomes of those who received only plasma with those who received PCCs (with or without plasma) for management of coagulopathy in patients who underwent cardiac surgery with cardiopulmonary bypass at a single institution from 2012 to 2016. Propensity score matching was used to obtain between-group balance. Primary outcome was avoidance of perioperative red cell transfusions. Other outcomes were incidence of massive transfusion (more than nine red cell units), refractory bleeding (requiring factor VIIa), and adverse events. RESULTS: Of 6362 patients, 1151 (18.2%) received plasma without any PCCs, and 204 (3.2%) received PCCs, either with (n=125) or without plasma (n=79). Overall, patient risk-profile was higher in the PCCs group. In a well-balanced propensity score match that included 117 patients per group, the odds ratio (OR) for red cell avoidance was 2.4-fold [95% confidence interval (CI) 1.2-4.8] higher in the PCCs group. Massive transfusion (OR 0.58; 95% CI 0.33-1.0) and refractory bleeding (OR 0.49; 95% CI 0.24-1.03) incidences were almost significantly lower in the PCCs group. The adverse event profiles were similar. CONCLUSIONS: Our exploratory study suggests that the use of PCCs as part of a multifaceted coagulation management strategy may have blood-sparing effects. Their incorporation into clinical practice, however, must await determination of their risk-benefit profile via multicentre randomised trials.

Greater intravenous fluid volumes are associated with prolonged recovery after colorectal surgery: a retrospective cohort study.

BACKGROUND: We carried out a retrospective assessment of whether perioperative fluid volume was associated with length of hospital stay (LOS) after colorectal surgery. METHODS: A single-centre chart review was conducted on colorectal surgeries that took place between January 2008 and December 2013. The primary outcome was LOS, with prolonged LOS defined as greater than median LOS. Secondary outcomes included postoperative pulmonary oedema, acute renal failure, myocardial infarction, and mortality. Univariate analysis, multivariable logistic regression, and quantile regression analyses were conducted to examine the association between perioperative fluid volume and prolonged LOS. RESULTS: Of the 1242 procedures, 57% were elective, 62% oncological, and 31% laparoscopic. The median LOS was 8.2 days (interquartile range 5.2, 14.7). Patients received 3.2 (sd 1.5) litres of fluid in the perioperative period (operating and recovery rooms), predominantly crystalloid. The volume (in litres) of perioperative fluid was independently associated with prolonged LOS (odds ratio 1.23, 95% confidence interval 1.10-1.36, P<0.01). This association persisted across the spectrum of definitions for prolonged LOS (10th-90th percentile). Logistic regression analysis also revealed that prolonged LOS was associated with age >65 yr, Charlson Co-morbidity Index ≥3, use of colloids, emergent surgery, estimated blood loss >200 ml, preoperative anaemia, erythrocyte transfusion, open surgeries, and surgical duration >4 h (C-statistic=0.79, Hosmer-Lemeshow=0.36). CONCLUSIONS: Greater perioperative fluid volume was independently associated with prolonged duration of recovery across a spectrum of surgical risk profiles. Fluid restriction should be considered a part of the care package in enhanced recovery after surgery programmes for colorectal surgery.

Red cell transfusion and the immune system.

Understanding the complex immunological consequences of red cell transfusion is essential if we are to use this valuable resource wisely and safely. The decision to transfuse red cells should be made after serious considerations of the associated risks and benefits. Immunological risks of transfusion include major incompatibility reactions and transfusion-related acute lung injury, while other immunological insults such as transfusion-related immunomodulation are relatively underappreciated. Red cell transfusions should be acknowledged as immunological exposures, with consequences weighed against expected benefits. This article reviews immunological consequences and the emerging evidence that may inform risk-benefit considerations in clinical practice.

Vascular targeted photodynamic therapy with palladium-bacteriopheophorbide photosensitizer for recurrent prostate cancer following definitive radiation therapy: assessment of safety and treatment response.

PURPOSE: Tookad is a novel intravascular photosensitizer. When activated by 763 nm light, it destroys tumors by damaging their blood supply. It then clears rapidly from the circulatory system. To our knowledge we report the first application of Tookad vascular targeted photodynamic therapy in humans. We assessed the safety, pharmacokinetics and preliminary treatment response as a salvage procedure after external beam radiation therapy. MATERIALS AND METHODS: Patients received escalating drug doses of 0.1 to 2 mg/kg at a fixed light dose of 100 J/cm or escalated light doses of 230 and 360 J/cm at the 2 mg/kg dose. Four optical fibers were placed transperineally in the prostate, including 2 for light delivery and 2 for light dosimetry. Treatment response was assessed primarily by hypovascular lesion formation on contrast enhanced magnetic resonance imaging and transrectal ultrasound guided biopsies targeting areas of lesion formation and secondarily by serum prostate specific antigen changes. RESULTS: Tookad vascular targeted photodynamic therapy was technically feasible. The plasma drug concentration was negligible by 2 hours after infusion. In the drug escalation arm 3 of 6 patients responded, as seen on magnetic resonance imaging, including 1 at 1 mg/kg and 2 at 2 mg/kg. The light dose escalation demonstrated an increasing volume of effect with 2 of 3 patients in the first light escalation cohort responding and all 6 responding at the highest light dose with lesions encompassing up to 70% of the peripheral zone. There were no serious adverse events, and continence and potency were maintained. CONCLUSIONS: Tookad vascular targeted photodynamic therapy salvage therapy is safe and well tolerated. Lesion formation is strongly drug and light dose dependent. Early histological and magnetic resonance imaging responses highlight the clinical potential of Tookad vascular targeted photodynamic therapy to manage post-external beam radiation therapy recurrence.

The effects of a treatment protocol for cardiac surgical patients with excessive blood loss on clinical outcomes.

BACKGROUND AND OBJECTIVES: Excessive blood loss (EBL) is a common complication of cardiac surgery that is associated with adverse events. The objective of this before/after study was to determine whether the implementation of a protocol for management of cardiac surgical patients with EBL was associated with improved clinical outcomes. MATERIALS AND METHODS: In November 2002, a protocol for prompt identification and aggressive management of cardiac surgical patients with EBL was implemented at our institution. The independent relationship between protocol implementation and adverse outcomes was measured by comparing the outcomes of patients who received > or = 4 RBC (red blood cell) units within 1 day of surgery and were operated on before protocol implementation (2000-02) with those operated on after protocol implementation (2003-05), using multivariable logistic regression analysis to control for the effects of confounders. The primary outcome was a composite of adverse events that included death, renal failure, stroke, and sepsis. Bootstrapping was used to confirm the validity of the results. RESULTS: Of the 11,314 patients who underwent surgery during the study period, 1875 (16.6%) received > or = 4 RBC units within 1 day of surgery, with 958 and 917 in the pre- and postprotocol periods, respectively. The composite adverse outcome occurred in 164 (17.1%) patients in the preprotocol period and 115 (12.5%) patients in the postprotocol period (P = 0.005). Protocol implementation was independently associated with reduced odds of the composite adverse outcome (odds ratio 0.67; 95% confidence interval 0.50, 0.91; P = 0.01). This estimate was stable in bootstrap sampling. CONCLUSION: Implementation of a protocol to manage EBL in cardiac surgery was independently associated with improved outcomes.

A multivariable model for predicting the need for blood transfusion in patients undergoing first-time elective coronary bypass graft surgery.

BACKGROUND: The incidence of blood transfusion in coronary artery bypass graft (CABG) surgery remains high. Preoperative identification of those at high risk for requiring blood will allow for the cost-effective use of some blood conservation modalities. Multivariable analysis techniques were used in this study to develop a prediction rule for such a purpose. STUDY DESIGN AND METHODS: Data were prospectively collected for all patients undergoing elective first-time CABG surgery from January 1997 to September 1998 at a tertiary-care teaching hospital (n = 1007). The prediction rule was developed on the first two-thirds of the sample by using logistic regression methods to examine the relationship of patient demographics, comorbidities, and preoperative Hb with perioperative blood transfusion. The remaining one-third of the sample was used to validate the rule. RESULTS: The transfusion rate was 29.4 percent. The prediction rule included preoperative Hb (g/dL, OR 0.928, p<0.0001), weight (kg, OR 0.938, p<0.0001), age (years, OR 1.037, p<0.01), and sex (male/female, OR 0.493, p<0.01); receiver operating characteristic = 0.86. When externally validated, the rule had a sensitivity of 82.1 percent and a specificity of 63.6 percent (at a selected probability cutoff). CONCLUSION: A simple and valid prediction rule is developed for predicting the risk of blood transfusion in patients undergoing first-time elective CABG surgery.

Regulation of two members of the steroid-inducible cytochrome P450 subfamily (3A) in rats.

Monoclonal antibodies have been successfully isolated which are isozyme-specific for cytochrome P450p (3A1) or P4501 (3A2), two members of the steroid-inducible cytochrome P450 subfamily exhibiting 89% amino acid sequence homology, and these antibodies show less than 5% cross-reaction with 11 other cytochromes P450 (P450a-P450k). A library of 28 purified monoclonal antibodies was established and characterized as to epitope specificity. Appropriate antibodies were selected and utilized to investigate the regulation of expressed cytochrome P450p and P4501 proteins as a function of age, sex, and treatment of rats with various inducing agents. Cytochrome P450p is not detectable in hepatic microsomes from untreated immature or adult male and female rats. Following dexamethasone treatment, expression of cytochrome P450p is observed in all groups with the levels reaching 30-37% of total microsomal cytochrome P450. Administration of other inducers such as pregnenolone 16 alpha-carbonitrile also yield enhanced levels of cytochrome P450p. Measurable amounts of constitutive cytochrome P4501 were detected in hepatic microsomes from immature and adult males as well as immature females but not in adult females. Cytochrome P4501 expression is inducible by dexamethasone in immature rats of both sexes and adult males, although dexamethasone is more effective as an inducer of cytochrome P450p than cytochrome P4501. Hence, not only is cytochrome P4501 protein expressed in immature animals of both sexes, it is also inducible in both sexes. These studies show that constitutive expression and induction of steroid-inducible cytochrome P450s may vary as a function of age.

Inactivation of cytochrome P450 and inhibition of ferrochelatase by analogues of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine with 4-nonyl and 4-dodecyl substituents.

Cytochrome P450- and heme-destructive effects of the 4-nonyl and 4-dodecyl analogues of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) were determined using hepatic microsomal preparations obtained from untreated, beta-naphthoflavone-treated, and phenobarbital-treated chick embryos. The 4-nonyl analogue of DDC was less efficacious than 4-ethyl DDC and 4-hexyl DDC, but more efficacious than 4-dodecyl DDC with respect to cytochrome P450-destructive activity. In all hepatic microsomal preparations, cytochrome P450 destruction by 4-nonyl DDC was accompanied by loss of microsomal heme. In contrast, 4-dodecyl DDC caused loss of heme only in hepatic microsomal preparations obtained from phenobarbital-treated chick embryos. The ability of 4-nonyl DDC and 4-dodecyl DDC to lower ferrochelatase activity was compared with that of 4-ethyl DDC and 4-hexyl DDC in cultured chick embryo hepatocytes. As the length of the 4-alkyl group was increased, the ferrochelatase-lowering efficacy and potency of the DDC analogue decreased. The 4-dodecyl DDC analogue was unable to lower ferrochelatase activity, which accorded with the finding that the administration of 4-dodecyl DDC to phenobarbital-treated rats did not lead to the accumulation of an N-alkylprotoporphyrin. The ability of 4-nonyl DDC to lower ferrochelatase activity was attributed to the formation of N-nonylprotoporphyrin IX following the administration of 4-nonyl DDC to phenobarbital-treated rats.

Evidence for the stereoselective inhibition of chick embryo hepatic ferrochelatase by N-alkylated porphyrins.

3,5-Diethoxycarbonyl-1,4-dihydro-2,6-dimethyl-4-ethylpyridine and its 4-propyl analogue were administered to phenobarbital-pretreated rats. The N-alkylprotoporphyrins (N-alkylPPs) that were isolated from rat livers, viz., N-ethylPP and N-propylPP, were found to have greater ferrochelatase-inhibitory potency than the corresponding synthetic N-alkylPPs. The N-ethylPP that was isolated from rat liver was found to contain 72% of the NB plus NA regioisomers, whereas synthetic N-ethylPP contained 40% of the NB plus NA regioisomers. In contrast, the N-propylPP that was isolated from rat liver contained the same amount of the NB/A regioisomer(s) as synthetic N-propylPP (33%). The NB plus NA regioisomers of N-ethylPP and the NB/A regioisomer(s) of N-propylPP that were isolated from rat liver were found to be significantly more potent than the corresponding synthetic regioisomers. We conclude that 1) the ferrochelatase-inhibitory potency of N-ethylPP that is isolated from rat liver is greater than that of synthetic N-ethylPP, due to differences in both regioisomer composition and the inhibitory potency of the NB plus NA regioisomers and stereoisomers, and 2) the ferrochelatase-inhibitory potency of N-propylPP that is isolated from rat liver is greater than that of synthetic N-propylPP, due solely to the difference in the ferrochelatase-inhibitory potency of the NB/A regioisomer(s) and stereoisomers. From the enhanced ferrochelatase-inhibitory potency of the NB plus NA regioisomers of N-ethylPP and the NB/A regioisomer(s) of N-propylPP that were isolated from rat liver, relative to the corresponding synthetic N-alkyllPP regioisomers, it was inferred that 2- and 4-vinyl substituents located on pyrrole rings A and B contribute to the optimal binding of N-alkylPPs to the ferrochelatase active site.

Interaction of chemicals with cytochrome P-450: implications for the porphyrinogenicity of drugs.

Our studies lead us to suggest that in order for an organic chemical to cause porphyrin accumulation in chick embryo hepatocyte culture, it must be: 1. lipophilic; 2. resistant to biotransformation by phase I or phase II reactions to compounds which are devoid of biological activity; and 3. capable of interacting with cytochrome P-450 with concomitant destruction of the heme moiety or the generation of reactive oxygen species. The interaction of porphyrin-inducing chemicals with cytochrome P-450 results in lowering of a regulatory "free heme" pool. The "free heme" pool may be lowered by inhibiting one or more of the enzymes of heme biosynthesis or by destruction of the heme moiety of cytochrome P-450. Specific chemical features enable organic chemicals to inhibit ferrochelatase activity or to destroy the heme moiety of cytochrome P-450. Chemicals with a wide variety of structures are able to inhibit uroporphyrinogen decarboxylase activity.

Regulation of heme biosynthesis in chick embryo liver cells.

According to current evidence heme controls the heme biosynthetic pathway primarily by controlling translocation of inactive pre-ALA-S from the cytosol into the mitochondrion, where ALA-S is active. A secondary mechanism involves inhibition by heme of transcription of the ALA-S gene. Porphyrinogenic drugs act by lowering a regulatory "free heme pool" by three different mechanisms: (a) by mechanism-based inactivation of cytochrome P-450 resulting in N-alkylprotoporphyrin formation and ferrochelatase inhibition, (b) by mechanism-based inactivation of cytochrome P-450 resulting in continuous heme destruction, (c) by enhanced generation of active oxygen species which interact with an endogenous substrate to form an inhibitor of uroporphyrinogen decarboxylase. It is also possible that porphyrinogenic drugs may exert a direct effect on the nucleus to increase formation of ALA-S mRNA.

Disruption of hepatic heme biosynthesis after interaction of xenobiotics with cytochrome P-450.

Heme biosynthesis in hepatocytes is controlled by a free heme pool, which regulates delta-aminolevulinic acid synthase. Porphyrinogenic chemicals deplete the regulatory free heme pool by interacting with cytochrome P-450 thereby inhibiting heme biosynthesis and/or causing heme breakdown. Recent developments allow us to predict which groups of chemicals are likely to be porphyrinogenic. One group is exemplified by 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine. Heterocyclic compounds of this type cause mechanism-based inactivation of cytochrome P-450, leading to the formation of N-alkylporphyrins, with ferrochelatase-inhibitory activity resulting in lowering the free heme pool. Allylisopropylacetamide exemplifies a second group. Such compounds containing a terminal olefinic or acetylenic group, cause mechanism-based inactivation of cytochrome P-450. In the process, the heme moiety of cytochrome P-450 is destroyed and the free heme pool is lowered. A third group is exemplified by planar polyhalogenated or polycyclic aromatic hydrocarbons. These compounds induce specific cytochrome P-450 isozymes but are poor substrates. Active oxygen is formed, which interacts with a hepatic substrate to form a uroporphyrinogen decarboxylase inhibitor. Inhibition of this enzyme leads to depletion of the free heme pool.

Differential inhibition of hepatic ferrochelatase by regioisomers of N-butyl-, N-pentyl-, N-hexyl-, and N-isobutylprotoporphyrin IX.

A series of analogues of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC), viz. 4-butyl DDC, 4-pentyl DDC, and 4-hexyl DDC was administered to phenobarbital-pretreated rats. The N-alkylprotoporphyrins (N-alkylPP) isolated from the rat livers were separated into regioisomers by means of high performance liquid chromatography; the NB or NA (NB/A) regioisomers constituted 19-26% of the total regioisomers. Considerable ferrochelatase-inhibitory activity was found in the NB/A regioisomers; the NC or ND (NC/D) regioisomers had little ferrochelatase-inhibitory activity. These findings supported the idea that the ferrochelatase active site could accommodate alkyl groups larger than methyl only if they were present on the nitrogens of the A or B pyrrole rings of the N-alkylPP. The inactivity of 4-isobutyl DDC as a ferrochelatase-lowering agent was investigated. After injection of 4-isobutyl-DDC into phenobarbital-pretreated rats, N-isobutylPP was isolated from the rat livers and separated into its regioisomers; the NB/A regioisomer constituted 3.8% of the total regioisomers. The NB/A regioisomer was found to have appreciable ferrochelatase-inhibitory activity whereas the NC/D regioisomer was inactive. The inactivity of 4-isobutyl-DDC as a ferrochelatase-lowering agent was attributed to the small amount of NB/A regioisomer present in the N-isobutylPP regioisomer mixture.

Isolation of an N-alkylprotoporphyrin IX from chick embryo livers following the administration of 3,5-diethoxycarbonyl-1,4-dihydro-4-ethyl-2,6-dimethylpyridine.

The ferrochelatase-lowering activity of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) analogues in chick embryo hepatocyte culture has been assumed to be due to the formation of an N-alkylprotoporphyrin IX. This assumption required confirmation. For this reason the 4-ethyl analogue of DDC was administered to phenobarbital-pretreated 19-day-old chick embryos. This resulted in hepatic accumulation of a green pigment with ferrochelatase-inhibitory activity. The green pigment was identified as an N-alkylprotoporphyrin IX by comparison of the electronic absorption spectra of its dimethyl ester and Zn complex with the corresponding spectra obtained from synthetic N-ethylprotoporphyrin IX.

Ferrochelatase-inhibitory activity and N-alkylprotoporphyrin formation with analogues of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) containing extended 4-alkyl groups: implications for the active site of ferrochelatase.

The ferrochelatase-inhibitory activity, porphyrin-inducing activity, and cytochrome P-450- and heme-destructive effects of a variety of analogues of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) were studied in chick embryo liver cells. The ferrochelatase-inhibitory activity of the 4-butyl, 4-pentyl, 4-hexyl, and 4-cyclopropylmethyl analogues of DDC was considered to be due to the formation of the corresponding N-alkylporphyrins. These N-alkylporphyrins were isolated from the livers of phenobarbital-pretreated rats following administration of the corresponding DDC analogues. The 4-isobutyl analogue did not have ferrochelatase-inhibitory activity despite its ability to cause formation of an N-isobutylporphyrin in rat liver. The 4-chloromethyl analogue of DDC inhibited ferrochelatase activity. The inability to isolate an N-alkylporphyrin from rat liver with this analogue may be due to its lability. The porphyrin-inducing activity of these analogues depended on their ferrochelatase-inhibitory potency and lipophilicity. The DDC analogues caused cytochrome P-450 and heme destruction. The relative ferrochelatase-inhibitory activity of the DDC analogues has implications for a postulated model of the binding of porphyrins in the ferrochelatase active site.

Properties of 17- to 19-day-old chick embryo liver microsomes. Induction of cytochrome P-450, effect of storage at low temperature, and resistance to lipid peroxidation.

Maximal hepatic cytochrome P-450 levels were induced in the 17-day-old chick embryo (four to five times control) with a dose of sodium phenobarbital of 6 mg/egg/day for 2 days. Similar levels of hepatic cytochrome P-450 were induced with a dose of propylisopropylacetamide of 4 mg/egg/day for 1 day. Chick embryo hepatic microsomes from phenobarbital-pretreated or from untreated chick embryos could be stored for periods of 14 days at -70 degrees C without a decrease in cytochrome P-450 levels. Moreover, no significant differences was discerned between the degree of suicidal inactivation of chick embryo hepatic cytochrome P-450 by 3,5-diethoxycarbonyl-1,4-dihydro-2,6-dimethyl-4-ethylpyridine in fresh and stored microsomes. Unlike rat hepatic microsomes, chick embryo hepatic microsomes do not undergo lipid peroxidative loss of cytochrome P-450 and heme when incubated in the presence of NADPH.

The effects of dihydropyridine calcium antagonists on heme biosynthesis in chick embryo liver cell culture.

A variety of 1,4-dihydropyridine calcium antagonists were tested for porphyrinogenic activity in chick embryo liver cell culture. 3,5-Dimethoxycarbonyl-1,4-dihydro-2, 6-dimethyl-4-(ortho-nitrophenyl)pyridine (nifedipine) was shown to be a potent porphyrinogenic agent. This activity was shared by a number of related analogues, viz., the 4-phenyl, 4-(meta-nitrophenyl), 4-(para-nitrophenyl), 4-(ortho-methoxyphenyl), 4-(meta-trifluoromethylphenyl), and 4-(para-trifluoromethylphenyl) analogues and nitrendipine; nicardipine exhibited very weak activity. The porphyrinogenic potency of the 1,4-dihydropyridines did not parallel their calcium antagonist activity. Nifedipine did not exhibit ferrochelatase-lowering activity in chick embryo liver cell culture and uroporphyrin and heptacarboxylic acid porphyrin were the major porphyrins to accumulate. Nifedipine did not cause suicidal destruction of cytochrome P-450 in chick embryo hepatic microsomes. Because nifedipine possesses comparable porphyrinogenic activity to sodium secobarbital in chick embryo liver cell culture, caution is required if nifedipine or related drugs are administered to patients with hereditary hepatic porphyria.

Comparison of the effects of 3-ethoxycarbonyl-1,4-dihydro-2,4-dimethylpyridine and 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine on ferrochelatase activity and heme biosynthesis in chick embryo liver cells in culture.

3-Ethoxycarbonyl-1,4-dihydro-2,4-dimethylpyridine (EDP) was shown to lack the ferrochelatase-lowering activity of 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine (DDC) in chick embryo liver cells in culture. This was attributed to the inability of EDP to cause destruction of the heme moiety of cytochrome P-450 with concomitant formation of N-methylprotoporphyrin IX. EDP was less potent as a porphyrinogenic agent than DDC and caused the accumulation of uroporphyrin, heptacarboxylic porphyrin, and coproporphyrin in contrast with DDC which caused primarily protoporphyrin to accumulate. The inactivity of EDP as a ferrochelatase-lowering agent and its low porphyrinogenic potency was explained, at least in part, by its rapid transformation in aqueous solution to other nondihydropyridine products. The two ethoxycarbonyl substituents of DDC are therefore essential for N-methylprotoporphyrin formation, ferrochelatase-lowering activity, and optimal porphyrin-inducing activity.