Outcome of Pregnancy in Women with Glomerular Diseases

Over the last 16 years the evolution of 24 pregnancies in 17 women with biopsy-proven glomerular disease was analyzed. The underlying renal histology was IgA nephropathy in 8 cases, lupus nephritis in 7, mesangiocapillary glomerulonephritis type I in 1, and focal segmental glomerulosclerosis in 1. All but 2 had normal renal function before conception and 3 were hypertensive. Fetal survival rate was 75%. There were 6 preterm deliveries (33.3%), 3 newborns small for gestational age (17%), 1 stillbirth, and 5 therapeutic abortions. The perinatal mortality was 5.5%. De novo hypertension occurred in 8 pregnancies (33.3%). In 11 pregnancies (46%) increased proteinuria was diagnosed and in 6 (25%) a decline in maternal renal function was recorded. Permanent impairment of renal function was seen in 2 women with renal insufficiency before conception. Maternal hypertension and renal function impairment were associated more frequently with obstetric complications. In conclusion, pregnancy is safe for normotensive mothers with glomerular diseases and normal renal function. Hypertension and impaired renal function at conception seem to carry increased risk for mothers and fetuses. Low-dose immunosuppressive treatment during pregnancy is not harmful for the fetus.     

Glomerular disease and pregnancy. A study of 123 pregnancies in patients with primary and secondary glomerular diseases

The clinical course of 123 pregnancies in 86 patients with biopsy-proven glomerular diseases have been studied. In 35 women the onset of nephropathy occurred during pregnancy. No complications were observed in more than half of the pregnancies. In the others, one third of the complications were obstetrical or fetal accidents, one third were renal manifestations (hypertension or deterioration of renal function) and one third were both causes. The lowest incidence of complications was observed in patients with membranous nephropathy and the highest in membranoproliferative glomerulonephritis patients. There were 6 spontaneous late abortion, 6 stillbirths and 5 neonatal deaths. 17 deliveries were preterm and 7 fetuses were small for gestational age. Hypertension appeared in 24 pregnancies, in 13 of which it was reversible and related to superimposed preeclampsia and in 11 it persisted after delivery (5 of these 11 pregnancies were in patients with IgA nephropathy). Renal function deteriorated in 10 cases during pregnancy. The deterioration was reversible in 6 and progressive in 4 (2 of whom had membranoproliferative glomerulonephritis). It is suggested that in most patients pregnancy does not change the natural history of glomerular disease.     

Influence of pregnancy on IgA nephropathy

In an attempt to clarify the influence of pregnancy on the natural course of IgA nephropathy, the courses of 79 pregnancies occurring in 47 patients with the disease were studied. These resulted in 3 artificial and 10 spontaneous abortions, and two pre-term and 64 full-term deliveries. Fifty four maternity passbooks were analyzed. In 22 pregnancies (40.7%) proteinuria was increased during the third trimester, and in 13 (76.5%) of 17 pregnancies receiving postpartum urinalysis, urinary protein was decreased to the level of the first trimester within one month after delivery. In two of the remaining four patients with a persistent increase in proteinuria, renal biopsy was carried out two months after delivery, revealing focal glomerular sclerotic lesions, in addition to mild mesangial proliferation compatible with IgA nephropathy. These findings indicated that increased urinary protein observed in the two pregnancies might be attributed to a complication of pre-eclamptic focal glomerular sclerosis rather than exacerbation of underlying IgA nephropathy. The glomerular filtration rate (GFR), examined in 27 patients both before and after pregnancy, was decreased in only two patients (7.4%), but these reductions appeared to go with the individual natural course. In 6 (15.0%) of 40 pregnancies, proteinuria was increased within one month after delivery, and one of them was diagnosed both clinically and pathologically as the acute exacerbation of IgA nephropathy. These data suggest that patients with IgA nephropathy might show transient acute exacerbation just after delivery rather than during pregnancy, and that even if such exacerbations occurred, pregnancy might have little influence on the natural course of the disease.     

Renal disease in pregnancy. Three controversial areas: mesangial IgA nephropathy, focal glomerular sclerosis (focal and segmental hyalinosis and sclerosis), and reflux nephropathy

Whether gestation has adverse effects on the course of renal disease is controversial. Three diseases regarding which conflicting opinions are especially noteworthy are IgA nephropathy, focal and segmental hyalinosis and sclerosis, and reflux nephropathy. We analyzed 102 pregnancies in 65 women with IgA nephropathy, noting hypertension in 63% of the gestations (severe in 18%), decreases in renal function in 22%, and biopsy evidence demonstrating a significantly greater amount of glomerular proliferation and crescents as well as focal and segmental hyalinosis and sclerosis when the renal histology from women who were pregnant and those who had never conceived were compared. Our experience with glomerular sclerosis is limited to 28 gestations in 15 patients, but the clinical findings resemble those of women with IgA nephropathy who conceived. Finally, we analyzed 227 pregnancies in 95 women with normal renal function and reflux nephropathy comparing results to 118 gestations in 42 patients with evidence of dysfunction (serum creatinine SCr greater than 1.25 mg/dL). Women with preserved function had good outcomes in general, whereas hypertension (36%) and an accelerated decline in function (8%) were observed in the groups that had moderate renal insufficiency in the initial stage of pregnancy. Increased proteinuria was the best predictor of progression of reflux nephropathy in both groups.     

Influences of pregnancy on the natural course of chronic glomerulonephritis with impaired renal function]

In an attempt to clarify the influence of pregnancy on the natural course of the chronic glomerulonephritis with impaired renal function (glomerular filtration rate: GFR less than or equal to 70 ml/min), the courses of 14 pregnancies occurring in 10 patients (seven with IgA nephropathy, one with membranoproliferative glomerulonephritis, one with membranous nephropathy and one with hereditary nephropathy) were studied. In 8 patients GFR measured before pregnancies ranged from 46 to 70 ml/min and in the other two creatinine clearance estimated in the first trimester of pregnancies was 62 and 49 ml/min, respectively. The pregnancies resulted in 10 live births, one spontaneous abortion, one artificial abortion and 2 neonatal deaths. In 2 out of 10 live births fetal weight was less than 2500 g. In 3 of 11 pregnancies there was neither increase in urinary protein nor elevation of blood pressure during pregnancies, while seven (64%) had increased proteinuria during the third trimester, and 4 of them were also complicated with hypertension. In 6 of 10 patients, there was no decrease in GFR during pregnancies. In three patients GFR was decreased from 70 to 36 ml/min, 70 to 58 ml/min and 62 to 48 ml/min, respectively. However, these reductions were considered to go with the natural course of respective patients because the reduction slopes were almost the same or rather mild in comparison with those estimated before or after pregnancies. The other patient also had a transient increase in serum creatinine level during two pregnancies, but the reciprocals of serum creatinine concentration before and after the pregnanciesdeclined linearly with time. These data suggest that pregnancy might have little influence on the natural course of the chronic glomerulonephritis even if complicated with renal functional impairment defined as GFR of 70 ml/min or less.     

Proteinuria in 3 sequential pregnancies following a fourth renal transplant

Pregnancy posttransplantation, particularly after kidney transplantation, is becoming common. It poses a challenge for transplant physicians, obstetricians and neonatologists due to the possible adverse maternal and foetal outcomes. The available experience on multiple pregnancies posttransplantation is limited. This case study reports 3 successful pregnancies - 5, 13 and 20 years after fourth renal transplantation resulting in vaginal deliveries at 37, 34 and 38 weeks - in a patient with reflux nephropathy. She developed hypertension, proteinuria and abnormal renal function during gestation with each pregnancy, all of which reversed after delivery. The reported case demonstrates successful foetal outcomes and reversible proteinuria, hypertension and allograft dysfunction possibly related to preeclampsia in the mother during her 3 successful pregnancies after a fourth renal transplant.     

Pregnancy in IgA nephropathy, reflux nephropathy, and focal glomerular sclerosis

Fetal outcome was retrospectively studied in 217 pregnancies observed during the past two decades in 93 patients, 34 suffering from IgA nephropathy (IgAGN, 69 pregnancies), 53 from reflux nephropathy (RN, 137 pregnancies), and six from focal glomerular sclerosis (FGS, 10 pregnancies). Overall incidence of live births was 175 in 217 (81%). Fetal loss, corrected for induced abortions, was 10 in 66 (15%) in IgAGN, 18 in 129 (14%) in RN, and 2 in 10 in FGS. Renal failure and hypertension preexisting prior to conception or developing early in pregnancy were the most important factors associated with unsuccessful fetal outcome whereas urinary tract infection had limited effects in RN patients. Influence of pregnancy on the course of maternal renal disease was evaluated in the same groups of patients. An abnormally rapid deterioration of renal function was observed in three of the women with IgAGN and in one of the RN patients (with an additional case among 46 further female RN patients) but in none in the FGS group. All five women experiencing functional deterioration had a serum creatinine (SCr) level of greater than or equal to 200 mumol/L (2.3 mg/dL) and hypertension at conception. Hypertension in pregnancy was highly predictive of recurrence of hypertension in subsequent pregnancy and of the remote development of permanent hypertension in IgAGN patients. We conclude that when renal function is preserved, pregnancy is usually successful and no deleterious effects on maternal renal disease are to be expected in patients with IgAN, RN, and probably FGS.(ABSTRACT TRUNCATED AT 250 WORDS)     

Renal survival rate of IgA nephropathy

In an attempt to identify prognostic indicators in IgA nephropathy, we evaluated the relationship between clinical and histological findings and changes in renal function in 81 patients with IgA nephropathy whose creatinine clearance was more than 80 ml/min at the time of renal biopsy. The incidence of patients whose creatinine clearance decreased to less than 60 ml/min during the follow-up period was calculated with the life table method to designate the renal survival rate. This rate was compared according to the clinical and histological findings at the time of renal biopsy. In conclusion, a statistically significant decrease in the renal survival rate was observed in patients with proteinuria of more than 1.0 g/day, hypertension, severe diffuse proliferative glomerulonephritis, diffuse proliferative glomerulonephritis with focal crescents and glomerular deposition of IgM and/or fibrinogen-related antigen.     

Thin basement membrane nephropathy in pregnancy

There are several published series of pregnancy in patients with nonimmunoglobulin A mesangial proliferative glomerulonephritis (most of whom have thin basement membrane nephropathy [TBMN]). The aim of the present study was to review the maternal and fetal outcomes of pregnancy in women with TBMN. The medical and obstetric histories of 86 women with TBMN and their 182 pregnancies (one twin) were reviewed. Data were collected retrospectively in 164 pregnancies (90%) and prospectively in 18 pregnancies (10%). Hypertension (alone or with proteinuria) developed in 15 unmonitored pregnancies (9%), and proteinuria alone developed in the third trimester in 2 pregnancies (1%). Hypertension was more common in the prospectively monitored pregnancies (6 pregnancies, 33%). In all, there were 174 live births (95%), and all fetal deaths occurred in the first and second trimesters in the absence of maternal complications. However, all the mothers of the 4 small for gestational age babies had been hypertensive. In TBMN, maternal hypertension, prematurity, and small for gestational age rates did not exceed those in the normal population. Overall, pregnancy in women with TBMN does not appear to be attended by a significantly increased maternal or fetal risk.     

Specific controversies concerning the natural history of renal disease in pregnancy

Whether or not pregnancy adversely affects the natural course of underlying primary renal disease, and whether fetal outcome is influenced by the type of renal disease per se are controversial issues. We retrospectively analyzed the fetal and maternal outcome in 148 women with various, biopsy-proven histological types of primary chronic glomerulonephritis (GN), including IgA GN (52 patients), membranous GN ([MGN] 20 patients), membranoproliferative type 1 GN ([MPGN] 58 patients), focal and segmental glomerulosclerosis ([FSGS] 13 patients), and minimal change nephrotic syndrome ([MCNS] 22 patients), who were pregnant (with a total of 290 pregnancies) after the clinical onset of GN, and in 104 women with reflux nephropathy (with a total of 254 pregnancies). Fetal outcome was poor in the presence of uncontrolled hypertension, nephrotic range proteinuria, and/or impaired renal function at conception or early in gestation, whatever the type of renal disease. An accelerated, more rapid than expected, worsening of maternal renal function was observed in five GN patients of whom four (two IgA, two MPGN) had serum creatinine (Scr) levels greater than 160 mumol/L (1.8 mg/dL) early in gestation, and in five patients with reflux nephropathy whose Scr at conception ranged from 180 to 490 mumol/L (2.0 to 5.5 mg/dL).(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of gestation on renal function in gravida with IgA nephropathy

Nineteen pregnancies in 17 women with IgA nephropathy (IgAGN) were studied in terms of the influence of gestation on the natural course of renal function in IgAGN. We performed serial examinations of the serum creatinine (S-Cr) levels before and during pregnancy and after delivery. Group I comprised 11 pregnancies in 10 gravida who revealed proteinuria (1.1 +/- 0.7 g/day; mean +/- SD) and microhematuria before pregnancy. The S-Cr before pregnancy averaged 0.86 +/- 0.13 mg/dl. Group II comprised 8 pregnancies in 7 gravida who showed isolated microhematuria. The S-Cr before pregnancy averaged 0.81 +/- 0.06 mg/dl. In group I, S-Cr did not decrease during pregnancy and was elevated at delivery and at 1-4 weeks after delivery (1.03 +/- 0.29 mg/dl, p less than 0.05) in comparison with the level before pregnancy. In group II, S-Cr decreased significantly during pregnancy and was not elevated at delivery or after delivery. The above results suggest that gestation had a slight and transient adverse effect on renal function in IgAGN with proteinuria.     

Thin basement membrane nephropathy associated with other glomerular diseases

Many reports confirm that thin basement membrane nephropathy (TBMN) commonly occurs together with other glomerular diseases such as minimal change glomerulonephritis, diabetes, membranous nephropathy, immunoglobulin (Ig)A glomerulonephritis, and focal segmental glomerulosclerosis. We postulate 3 explanations for these observations. The association of minimal change glomerulonephritis with TBMN probably is artifactual whereas the association with diabetes and membranous glomerulonephritis probably is coincidental. However, the link between TBMN and IgA disease and focal segmental glomerulosclerosis may be pathogenetic. Clinical evidence indicates that the presence of an associated glomerulopathy significantly worsens the prognosis of TBMN. Thus, patients with TBMN and another glomerular lesion usually have more marked proteinuria, and are more likely to have hypertension and renal insufficiency. The frequency of another glomerulopathy in patients with TBMN means that all patients in whom TBMN is suspected but who have heavy proteinuria or renal insufficiency should undergo a renal biopsy examination. However, there is no evidence that TBMN alters the prognosis of another glomerulopathy, and, in particular, patients with TBMN and IgA disease do not have different clinical features or a worse prognosis than those with IgA disease alone.     

Pregnancy after donor nephrectomy

The use of living-related kidney donors has been a routine practice in most major transplant centers in the United States for more than 20 years. Concern has arisen regarding the potential for developing hypertension and progressive renal dysfunction after renal donation. Pregnancy results in hyperfiltration and might be an added risk for the development of hypertension, proteinuria, or renal insufficiency in donors. From 1963 until 1984, the Cleveland Clinic Foundation performed 1031 renal transplants, 355 from living donors. Of these 355 living donors, 191 were female, and of these, 23 successfully conceived after kidney donation. Prenatal and delivery records of all 23 were reviewed. There were 39 pregnancies in 23 women with 32 viable births. Conception ranged from 2 weeks to more than 9 years postnephrectomy. Mean blood pressure at the time of donor evaluation was 120/75 mm Hg (SD: +/- 11/8 mm Hg). Mean blood pressure during pregnancy was 114/68 mm Hg (SD: +/- 7/6 mm Hg). One plus proteinuria was detected in 2 women during the third trimester and trace proteinuria was seen in 7 pregnancies; this proteinuria disappeared after delivery. Thirteen of twenty women who carried to term were reevaluated 2-14 years after donor nephrectomy. All parameters studied were normal. Mean length of follow-up after donor nephrectomy was 7.9 years. These data suggest that, after donor nephrectomy, women can have a normal pregnancy without significant problems related to the kidney donation. Also, hyperfiltration associated with the combination of unilateral nephrectomy and pregnancy does not lead to significant hypertension, proteinuria, change in glomerular filtration rate, or abnormalities of the urinary sediment.     

Consecutive Spontaneous Triplet and Twin Pregnancies in a Woman After Renal Transplantation

BackgroundThere are numerous reports of successful pregnancies following kidney transplantation. However, little information is available regarding the management and evolution of multiple pregnancies in a kidney-transplanted woman.Case reportWe report the case of successful consecutive spontaneous triplet and twin pregnancies in a woman who had undergone kidney transplantation at 30 years of age, 12 years before the first pregnancy, as a result of end-stage renal disease secondary to chronic glomerulonephritis due to diffuse proliferative lupus nephritis. An integrated multidisciplinary team closely followed progress during the pregnancies. Maternal complications during the pregnancies included light proteinuria, controlled hypertension, and anemia. No graft rejection episodes or deterioration of renal function was noted during the pregnancies or after the deliveries.ConclusionCurrently, more than 2 years after her last pregnancy, the mother and all 5 babies are healthy and the mother’s renal transplant function is normal.     

Long-term beneficial effects of angiotensin-converting enzyme inhibition in patients with nephrotic proteinuria.

Angiotensin-converting enzyme inhibitors (ACEI) can reduce proteinuria in diabetic and nondiabetic nephropathy. However, no studies have determined whether this antiproteinuric effect modifies the progression of renal insufficiency. We studied the evolution of 46 nondiabetic patients with nephrotic proteinuria treated with captopril for a minimum of 12 months. The follow-up period before captopril treatment was 12 to 18 months. At the end of follow-up, after captopril introduction (24.4 +/- 7.6 months), proteinuria had decreased from 6.3 +/- 2.5 to 3.9 +/- 3.1 g/24 h (P less than 0.001), with a mean decrease of 45% +/- 28%. The proteinuria decrease was higher in patients with reflux nephropathy, proteinuria associated with reduction of renal mass, inactive crescentic glomerulonephritis, nephroangiosclerosis, and IgA nephropathy, whereas patients with membranous glomerulonephritis and idiopathic focal glomerulosclerosis showed a poorer response. Patients were separated according to a proteinuria reduction greater (group A, 23 patients) or lower (group B, 23 patients) than 45% of the initial value. At the end of follow-up, renal function had not significantly changed in group A with respect to values at the start of treatment: serum creatinine (SCr) was 229 +/- 167 mumol/L (2.6 +/- 1.9 mg/dL) versus 203 +/- 97 mumol/L (2.3 +/- 1.1 mg/dL), and creatinine clearance (CrCl) was 0.80 +/- 0.52 mL/s (48 +/- 31 mL/min) versus 0.87 +/- 0.47 mL/s (52 +/- 28 mL/min). The slope of the reciprocal of Scr (1/SCr) showed a significantly beneficial change after captopril introduction.(ABSTRACT TRUNCATED AT 250 WORDS)     

Paulista Registry of glomerulonephritis: 5-year data report.

BACKGROUND: The Paulista Registry of Glomerulopathies was created in May 1999 and comprises several centres of S?o Paulo, the most populous Brazilian State, that concentrates people from all regions of the country who look for health care. METHODS: This report includes data from 2086 patients from Brazil submitted to renal biopsy due to the presumed diagnosis of glomerular diseases, registered prospectively since May 1999 until January 2005. Data were collected by the integrants of the 11 centres involved, utilizing a standardized questionnaire. RESULTS: The mean age of the patients was 34.5+/-14.6 years. Primary glomerular diseases were more frequent in males (55.1%) than in females; on the other hand, secondary glomerular diseases were more frequent in females (71.8%). The most common clinical presentation was nephrotic syndrome and the frequency of hypertension, at this time, was 55.5%. There was a predominance of indication of biopsies in the third, fourth and fifth decades of life. The most common primary glomerular diseases were focal and segmental glomerulosclerosis (29.7%), followed by membranous nephropathy (20.7%), IgA nephropathy (17.8%), minimal change disease (9.1%), membranoproliferative glomerulonephritis (7%), crescentic glomerulonephritis (4.1%), advanced chronic glomerulopathy (4%), non-IgA mesangial glomerulonephritis (3.8%), diffuse proliferative glomerulonephritis (2.5%), focal segmental proliferative glomerulonephritis (1%) and others (0.3%). The most frequent secondary glomerular disease was lupus nephritis, corresponding to 66.2% of the cases, followed by post-infectious glomerulonephritis (12.5%), diabetic nephropathy (6.2%), diseases associated to paraproteinaemia (4.9%), hereditary diseases (4.6%), vasculitis (3.2%), malignancies (0.9.%), secondary focal segmental glomerulosclerosis (0.6%) and others (0.9%). CONCLUSION: Focal segmental glomerulosclerosis was the most frequent primary glomerular disease, followed by membranous nephropathy and IgA nephropathy. Lupus nephritis predominated over all the other secondary glomerular diseases.     

Systemic hypertension in patients with glomerulonephritis

Although systemic hypertension is very common in patients with glomerulonephritis there is a dispute if this alteration is consequence of the glomerulonephritis "per se" or is a consequence of the renal failure secondary to the glomerular lesion. With the aim to analyze the factors associated with systemic hypertension, 196 patients with different forms of nephritis were studied. The systemic arterial pressure was measured by standard sphygmomanometer, renal function was evaluated by the determination of the serum creatinine concentration or creatinine clearance. The diagnosis of the type of glomerulonephritis was made on the basis of an examination of kidney biopsy specimens. The prevalence of arterial hypertension among patients with glomerulonephritis was 62.7%. The hypertensive patients were older (hypertensive = 30.6 +/- 12.8; normotensive = 25.4 +/- 1.6 years; P = 0.03). The prevalence of arterial hypertension was lower in patients with minimal glomerular lesion (12.5%), though their ages were also lower (18.1 +/- 3.6 and 29.1 +/- 1.03 years; P = 0.03). Arterial hypertension did not correlate with the serum levels of creatinine and albumin; creatinine clearance and 24-h proteinuria. IN CONCLUSION: In the patients with glomerulonephritis, the presence of arterial hypertension was associated with a higher mean age whereas the intensity of proteinuria, the level of renal function or the type of glomerulonephritis was not different between the two groups.     

Membranous glomerulonephritis and pregnancy

The clinical courses of 33 pregnancies in 24 patients with biopsy proven membranous glomerulonephritis have been analyzed. Twenty-four percent (8) of pregnancies resulted in fetal loss, 43% (14) in premature delivery and 33% (11) in a live birth after 36 weeks gestation. Maternal renal function declined during pregnancy in 9% (3) of the pregnancies and in 46% (15) hypertension developed. In 55% (18) proteinuria increased significantly during pregnancy. In 30% (10) nephrotic range proteinuria was recorded in the first trimester. Presence of nephrotic range proteinuria during the first trimester correlated with both poor fetal and poor maternal outcome (p less than 0.0004 and p less than 0.0002, respectively). It is concluded that pregnancy in patients with membranous glomerulonephritis is associated with increased fetal loss and, in some instances, a worsening in maternal renal function. The literature on this topic is reviewed in relation to these findings.     

An overview of pregnancy in women with underlying renal disease

This is a report of a retrospective study of the effects of preexisting glomerular disease and pregnancy on each other. Two hundred forty pregnancies in 166 Japanese women who delivered between 1970 and 1988 were analyzed. There were 206 (86%) live births, 14 (6%) perinatal deaths, and 20 (8%) spontaneous abortions. Perinatal loss was greatest in women with hypertension and/or glomerular filtration rate (GFR) less than 70 mL/min before conception. Pregnancy did not appear to adversely affect the underlying glomerular disease if GFR was greater than 70 mL/min and blood pressure was below 140/90 mm Hg. However, with moderately impaired renal function (creatinine greater than 124 mumol/L [1.4 mg/dL] or GFR less than 50 mL/min), the long-term prognosis was poorer, despite generally favorable obstetrical outcomes. Gravidas with membranoproliferative glomerulonephritis had the highest rates of hypertension (29%) and decreased renal function (33%) at final follow-up, ie, the type and severity of glomerulonephritis had a major impact on clinical course.     

Pregnancy in women with diffuse mesangial proliferative glomerulonephritis

The clinical course of 168 pregnancies in 91 women with non-IgA diffuse mesangial proliferative glomerulonephritis has been analyzed. Twenty percent (33) of pregnancies resulted in fetal loss, 18% (31) in premature delivery and 62% (105) in a term infant. Maternal renal function declined, reversibly, in 3% (5) of pregnancies and in 48% (80) hypertension developed. In 53% (89) a significant increase in proteinuria occurred in pregnancy. Fetal and maternal complications of pregnancy occurred more frequently in patients with pre-existing hypertension although differences failed to reach statistical significance (p greater than 0.01). The presence of severe vessel lesions on the diagnostic renal biopsy was associated with a significantly higher fetal loss and prematurity rate (p less than 0.0005 and p less than 0.005, respectively).