BRIEF REVIEWS Parathyroid Hormone-Related Protein
hormone. The cDNA-predicted amino acid sequence of PTHrP contains multi-
A Peptide of Diverse Physiologic Functions
ple basic motifs, suggesting that this peptide, like many neuroendocrine peptides, undergoes extensive posttranslational processing. The posttranslational processing of PTHrP is extremely complex and appears to be analogous to that of proopiomelanocortin (POMC). Like
Anne E. de Papp and Andrew F. Stewart
Parathyroid hormone-related protein (PTHrP) is the factor responsible for the syndrome of humoral hypercalcemia of malignancy. PTHrP is produced by a multitude of normal as well as malignant cells, and exerts both classic parathyroid hormone (PTH)-like and PTH-unlike effects. The molecular cloning of the PTHrP gene, and the subsequent recognition of its widespread expression in normal tissues under normal physiologic conditions, has prompted intense inquiry into its biologic function. PTHrP appears to act in an autocrine or paracrine fashion in (a) normal embryogenesis and neonatal development, (b) cellular growth and differentiation, (c) reproduction and lactation, (d) epithelial calcium transpovt, and (e) smooth muscle relaxation. These five key emerging physiologic roles of PTHrP are the focus of this review. (Trends Endomino1 Metab
1993;4:
18 l-l 87)
In 1941 Albright speculated that certain tumors might produce a hormonal sub-
first 13 amino acids) accounts for their common ability to stimulate adenylate
stance, similar to parathyroid hormone (PTH), that was responsible for the hypercalcemia complicating certain malignancies (Albright 194 1). Although malignancy-associated hypercalcemia was first recognized in the 192Os, it was not until
cyclase in both kidney and bone. The nowclassic biochemical stigmata of HHM are believed to be mediated through the recently cloned PTHlPTHrP receptor (Juppner et al. 199 1). The primary amino acid sequence of PTHrP is highly evolu-
1987 that the peptide responsible for humoral hypercalcemia of malignancy (HHM) was purified from tumor extracts and its cDNA molecularly cloned. The peptide, parathyroid hormone-related protein (PTHrP), was so named for its amino acid
tionarily conserved across species, suggesting that it plays a key role in normal cellular function. The PTH and PTHrP genes are thought to have arisen from a common ancestral gene, through an ancient chromosomal duplication event. The human
homologyandpathophysiologicmimicry of the actions of PTH. Patients with HHM, like those with hyperparathyroidism, display increased nephrogenous CAMP excretion, a reduced renal phosphate threshold and increased osteoclastic bone resorption. The N-terminal
homology
PTHrP gene, located on chromosome 12, is a complex gene consisting of eight exons, allowing for alternative splicing to yield three different mRNA transcripts. In humans, the PTHrP gene encodes three isofonns of 139,141, and 173 amino acids,
between
PTH and PTHrP (they share eight of the
whereas in the chicken two isoforms exist, and in the rat and mouse only a single mRNA species has been found.
Anne E. de Papp and Andrew F. Stewart are at the Division of Endocrinology, West Haven Veterans Affairs Medical Center, West Haven, CT 06516; and the Division of Endocrinology, Yale University School of Medicine, New Haven, CT 06510, USA.
TEM Vol.4,No. 6,1993
PTHrP appears to be a polyhormone with different physiologic functions that depend on the particular fragment secreted, which in turn is determined by tissue-specific processing of the pro-
01993,Elsevier
Science
Publishing
Co., 1043-2760/93/$6.00
POMC, which is cleaved intracellularly to yield a family of peptide hormones (ACTH, MSH, y-lipotropin, endorphins, and so on), PTHrP appears to be processed into a family of related-but-distinct mature peptides with potentially different physiologic functions. The history of HHM, the purification of PTHrP, the structure of the PTHrP gene, and the posttranslational processing of PTHrP have been the subject of a number of recent reviews (Halloran and Nissenson 1992, Stewart and Broadus 1990, Broadus and Stewart 1993, Strewler and Nissenson 1990, Stewart et al. 1993). Unlike PTH, which is normally expressed and secreted only by parathyroid tissue, PTHrP is expressed and secreted by a variety of normal as well as pathologic tissues (Table 1). The most abundant physiologic sources of PTHrP appear to be milk, lactating breast tissue, and the amnionic membrane. Although its role in HHM is now well accepted and reasonably well understood, the normal physiologic roles of PTHrP are only beginning to be explored. This review focuses on five major emerging roles for PTHrP in normal physiology: the role of PTHrP (a) in fetal growth and development; (b) as a cellular growth factor; (c) as a hormone and paracrine factor in gestation, reproduction, and lactation; (d) in transepithelial calcium transport; and e) as a widely expressed smooth muscle relaxant.
??
PTHrP in Fetal Growth and Development
Experimental evidence suggests that PTHrP plays a role in the normal growth and development of both embryonic and mature tissues, and may be a key factor in cellular differentiation. PTHrP is widely expressed in a spectrum of mammalian and avian fetal tissues, including those of
181
Table 1. Tissue survey of parathyroid gene expression Tissue
hormone-related
+ + _
Rat
Human
+ + +
+ +
Heart Kidney Liver
-
+ -
Lung
+
+
+ + + +
+ + +
+ + +
+
-t +
Mammary gland Ovary Parathyroid Placenta Prostate Skeletal muscle Skin Small intestine Spleen Stomach Testis Thymus Urinary bladder Uterus Vascular smooth muscle
+
t
+
t
+
+ + +
+ + + + + + +
t
t + t
t
+
f
+
t
t
The expression of PTHrP is either detectable (+) or absent/below the limit of detection (-) in these tissues. Blank spaces indicate that the tissue has not been tested. From Halloran and Nissenson (1992). with permission.
the human fetus, where it may regulate growth in utero (Halloran and Nissenson 1992). Immunohistochemical examination of human fetal tissue, obtained from ectopic and aborted pregnancies between 8 and 12 weeks of gestation, localizes
old neonatal rat, glomerular staining is completely absent, whereas tubular staining persists (Burton et al. 1992). Comparable studies on the musculoskeletal system have revealed changing patterns of staining during skeletogenesis. Immature
PTHrP to the fetal central and peripheral nervous systems, salivary and adrenal glands, pancreas, liver, lung, epidermis, oral and gastrointestinal epithelium, and throughout the urogenital and musculoskeletal systems (Moniz et al. 1990). Similar widespread expression of PTHrP has been described during chick and rat embryogenesis (Schermer et al. 199 1, Campos et al. 1991).
limb bud mesenchyme stains for PTHrP, but subsequently loses all staining once mature cartilage has formed (Moniz et al. 1990). Interestingly, immunoreactive PTHrP is found within fetal rat perichondrium one day before the onset of ossifi-
Developmental and temporal changes in the location and intensity of PTHrP gene expression appear to occur during embryogenesis. Northern blot analysis of human fetal tissue reveals multiple PTHrP mRNA transcripts displaying tissue-specific patterns of expression (Moniz et al. 1990). For example, the developing human fetal kidney shows a variable pattern of PTHrP immunostaining depending on gestational age. At 10 weeks’ gestation, PTHrP is present in the fetal glomeruli and renal tubules. By 25 weeks, however, glomerular staining is greatly reduced (Burton et al. 1992). In the 2-day-
182
phenotype.
Furthermore,
this differenti-
ated phenotype expresses both PTHrP and
Chicken
Bone Brain
protein (PTHrP)
cation (Heath et al. 1990), suggesting a possible role for PTHrP in fetal skeletal mineralization and limb bud formation. These observations support the concept that PTHrP has an important role in the embryologic development of mammalian
PTH/PTHrP receptor mRNA, suggesting that PTHrP may upregulate its own receptor. Lastly, “knockout” of the PTHrP gene in transgenic mice is lethal. Neonatal mice die shortly after birth and display a multitude of skeletal defects (Karaplis et al. 1992). This dramatically illustrates the critical importance of PTHrP in normal fetal growth and development.
??
FTHrP as a Cellular Growth Factor
PTHrP appears to function not only as a regulator of growth and development during embryogenesis, but may modulate normal adult cellular growth and differentiation as well. PTHrP functions as an autocrine growth factor or growth modulator in many cell lines, including renal carcinomas, lymphocytes, fibroblasts, osteoblasts, chondrocytes, and keratinocytes. For example, using both human, native PTHrP (purified from a breast carcinoma) and synthetic PTHrP ( l-36), Insogna et al. ( 1989) demonstrated that PTHrP possesses transforming growth factor (TGF)-like properties. In the presence of epidermal growth factor (EGF), PTHrP induces dose-dependent transformation of a rat kidney fibroblast cell line (NRK 49F) in soft agar. Furthermore, the authors found that, like TGFl3, PTHrP could stimulate fibronectin synthesis in human dermal fibroblasts. Further data support a role of PTHrP as a terminal differentiation factor in human keratinocytes (Holick et al. 1988). Since immunoreactive PTHrP can be detected in all layers of normal human epidermis (Atil-
and avian species.
lasoy et al. 199 1 ), PTHrP may play an autocrine or paracrine role in normal skin homeostasis via its TGF-like properties. A polyclonal PTHrP antiserum has been demonstrated to inhibit cell growth in a human renal cell carcinoma line (SKRC-
That PTHrP may play an additional role in the development and differentiation of the early preimplantation embryo is suggested by data from several investigators (Chan et al. 1990, van de Stolpe et al. 1993). F9 teratocarcinoma cells are a useful model of normal embryonic development, as they share many properties with pluripotent embryonic stem cells. The addition of hPTHrP (l-34) and retinoic acid to F9 cells promotes the differentiation of these cells into a parietal endodermlike
1) known to secrete PTHrP in vitro (Burton et al. 1990). In contrast, PTHrP (l-34) appears to function as an autocrine growth inhibitor in lymphocytes (Adachi et al. 1990). Similarly, Kaiser et al. (1992) showed a growth-inhibitory effect of amino-terminal PTHrP on a human keratinocytecell line. By infecting these cells with a full-length antisense cDNA sequence to PTHrP (and thereby blocking PTHrP synthesis and secretion), these authors observed a decrease in cell doubling
01993, Elsevier Science Publishing
Co., 1043-2760/93/$6.00
TEM Vol. 4, No. 6, 1993
time and an increase poration.
endogenous effective
in thymidine
One concludes,
incor-
therefore,
that
PTHrP can act as either an
inhibitor
growth, depending
or promoter
of cell
on the portion of the
molecule that is secreted and which target tissue is under consideration.
??
Table 2. The concentration of parathyroid hormone-related protein (PTHrP) in milk products
PTHrP as a Hormone of Reproduction
PTHrP appears to be tightly reproductive process in both and avian species. PTHrP is lactating mammary tissue, centa, oviduct, and the shell
PMP (l-34) amide, mean ng eq/ml f SD
5 6 6
50.0 + 14.9 95.6 + 34.0 75.7 f 42.8
Whole milk Low-fat milk
5 3
81.4 f 17.9 88.9 f 8.7
Nonfat milk Dried milk
3 3
118.3 + 18.9 76.2 f 32.7
Chocolate milk Buttermilk
3 3
6.8 f 6.3 4.5 + 4.2
Unprocessed
linked to the mammalian produced by uterus, plagland; in ad-
PTHrP in Lactation The physiologic role of PTHrP in lactation has been the subject of much speculation and study. Using Northern analysis, Thiede and Rodan (1988) showed that PTHrP is produced in rat lactating mammary tissue, but not in nonlactating mammary tissue. Furthermore, PTHrP expression appears to be a function of the suckling stimulus. Whereas PTHrP mRNA is undetectable in pregnant nonlactating breast tissue, it appears within 24 h postpartum with the onset of suckling, and high levels are maintained to at least 20 days postpartum. Time-course studies have revealed that PTHrP mRNA expression sharply decreases i-2 h after the removal of nursing pups and is absent by 4 h after the cessation of suckling; this effect can be reversed by the resumption of nursing (Thiede and Rodan 1988). Further investigation into the relationship between PRL and PTHrP has demonstrated that serum PRL, rather than suckling per se, controls the expression of PTHrP in mammary tissue. Bromocriptine, an inhibitor of PRL secretion, appears to inhibit PTHrP production by blocking the suckling-induced rise in PRL. Mammary tissue of nursing rats treated with bromocriptine has undetectable PTHrP mRNA levels. Furthermore, this effect can be overridden by the subcutaneous injection of ovine PRL (Thiede 1989). Although the data in rats are convincing, it remains questionable whether PRL controls PTHrP gene expression in other species. Khosla et al. (1990) found no correlation between serum PRL levels and serum PTHrP levels in normal lactating women or in seven women with pathol-
milk
Human Bovine Rat Commercial
dition, abundant quantities are found in both milk and amniotic fluid.
TEM Vol.4,No. 6,1993
n
Sample
Milk-based
milk products
infant formulas
Similac (Ross) Enfamil (Mead Johnson) SMA (Wyeth) Enfamil Premature 20 Similac 60/40 Enfamil Premature 24 Soy-based
infant formulas
3 3 3 3 3 3
32.8 f 6.8 * 5.9+ 7.3 * 1.1 &. 13.7 f
8.9 1.9 2.1 0.9 0.8 4.4
Undetectable
The results are expressed as nanogram equivalents of PTHrP per milliliter of milk: 1 ng eq/mL is equal to -0.25 nM. The concentration of PTHrP in plasma from patients with HHM, using these units, is -0.08 ng eq/mL. Thus, the values in milk are -lOOO-fold higher than in plasma of patients with HHM, and are -10,000. fold higher than in normals. From Budayr et al. (1989). with permission.
The extremely high levels of PTHrP found in milk (Table 2) [concentrations
trations of PTHrP in human milk that are 10,000 times higher than in normal serum. Similar elevations are found in both rat and bovine milk. Lower levels of PTHrP are present in various infant formulas and, interestingly, are undetectable in soy-based formulas (Budayr et al. 1989). The significance of the latter findings is uncertain, but it is interesting to note that soy-fed infants have increased 1,25-(OH), vitamin-D levels when compared with breast-fed infants (Hillman et al. 1988) and, further, formula-fed infants have an exaggerated fall in serum calcium immediately following birth as compared with breast-fed infants (Budayr et al. 1989). Normally, serum calcium levels transiently fall in term infants, with a nadir at 24-48 h after birth: one could propose therefore that maternal milk-derived
in human milk average 30,000-60,000 pM (Budayr et al. 1989)] in comparison to the levels reported in plasma of patients with HHM [the mean being 20 pM (Burtis et
PTHrP serves to blunt this early hypocalcemic response, perhaps allowing time for the maturation of the infant’s own parathyroid glands.
al. 1990)] suggest that PTHrP may play a critical role in mammary development and calcium transport during lactation and/or in neonatal development or calcium homeostasis. Budayr et al. (1989) and Burtis et al. ( 1990) reported concen-
Lactation in mammals, including humans, is accompanied by increased bone mobilization and net bone loss. This occurs independently of any changes in PTH, vitamin D, or calcitonin metabolism (Brommage and DeLuca 1985). In cows,
ogic hyperprolactinemia. Naturally, PTHrP content in breast tissue itself was not measured, nor was any correlation attempted with the PTHrP content of breast milk. Other factors appear to regulate PTHrP expression in mammary tissue, as increased PTHrP expression has been reported during late lactational stages in rats, a time when serum PRL levels are clearly declining (Yamamoto et al. 1992a). Finally, the injection of ovine PRL into preterm pregnant rats does not induce PTHrP expression in nonlactating mammary tissue (Thiede 1989), suggesting that other humoral agents that circulate during the postpartum period are required for the development of PRL-induced PTHrP expression.
01993, Elsevier Science Publishing
Co., 1043-2760/93/$6.00
183
may
reach sufficient
levels in the plasma
during lactation to exert a classic endocrine effect. Although these data are intriguing, the precise physiologic role of PTHrP during lactation remains to be determined.
tal unit. The role of PTHrP as a fetoplacental hormone is currently an area of active study in several laboratories.
PTHrP in Fetal Cakium
Homeostasis
Several observations support a role for PTHrP as the putative “fetal PTH.” There is little evidence that PTH itself is secreted in utero, nor does it appear to cross the
1. Diagrammatic representation of fetal and placental sources of parathyroid hormonerelated protein (PTHrP). Note that the fetal parathyroids also produce PTHrP. Adapted from Ferguson et al. (1992), with permission.
Figure
the calcium concentration
in milk is 1O-
15 times higher than that in blood, suggesting that an active calcium transport system exists between serum and milk (Law et al. 1991). PTHrP could potentially serve both a classic endocrine function (mediating bone resorption) and a paracrine or autocrine function (stimulating the transfer of calcium from maternal
blood
to milk) during lactation. Law et al. (1991) found a weakly positive correlation between the concentrations of calcium and PTHrP in milk from lactating cows. However, this observation
was not confirmed
by similar studies in lactating rats (Yamamoto et al. 1992a). Furthermore, the injection
of PTHrP-neutralizing
antisera
into lactating mice has no effect on calcium concentration
in either maternal serum or
milk, nor does it affect total body calcium in the rat pups (Yamamoto et al. 1992a). These latter observations fail to support the notion that PTHrP is a systemic calciummobilizing hormone in lactation. Furthermore, many studies of lactating women have failed to show detectable circulating plasma levels of PTHrP during lactation (Khosla et al. 1990, Budayr et al. 1989). One recent study, however, employing a different PTHrP immunoassay, has demonstrated elevated levels of serum PTHrP in twelve of 19 lactating women (range, 2.3-7.8 pmol/L) compared with undetectable levels in 16 bottle feeding, postpartum women (Grill et al. 1992). If confirmed, this would be evidence that PTHrP
184
by the finding noted above, that the cord blood of infants may contain measurable quantities of PTHrP. Thus, PTHrP may exert endocrine or paracrine effects on a host of target tissues within the fetoplacen-
placenta. Immunoreactive PTH is low or undetectable in human cord blood, yet PTH-like bioactivity is significantly increased compared with maternal levels (Allgrove et al. 1985). Rodda et al. (1988) have shown that the fetal parathyroid produces PTHrP and that 50% or more of the PTHlike bioactivity in the fetal parathyroid gland is derived from PTHrP as opposed to PTH. Furthermore, PTHrP may be elevated in the cord blood of infants. Hillman et al. (1990) reported that both term and premature infant cord blood contain 3 times higher concentrations of PTHrP than normal adult serum. Evidence reviewed below strongly suggests that fetal, as opposed to maternal, PTHrP plays a vital role in fetal-placental calcium transfer, and that this function is unique to a region in the PTHrP sequence not shared with PTH.
PTHrP and the Fetoplacental
Unit
In addition to the systemic maternal and neonatal effects of PTHrP described above, PTHrP appears to function as a hormone within the fetoplacental unit (Figure 1). PTHrP gene expression has been detected in human amnion, chorion, placenta, decidua, and myometrium; the most abundant signal is that of the amnionic membrane (Ferguson et al. 1992). PTHrP expression in the amnion rapidly decreases after the onset of labor and with rupture of amnionic membranes, suggesting that PTHrP downregulation may somehow play a permissive role in the onset of labor. Amnionic fluid itself contains meas urable quantities of PTHrP (40 pmol/L at term) (Ferguson et al. 1992). Thus, PTHrP could exert paracrine effects on fetal gastrointestinal epithelium, pulmonary epithelium, and skin, since the developing fetus swallows, inspires, and is bathed in amnionic fluid. Conversely, these tissues could be the source, in part, of amnionic fluid PTHrP. Evidence that PTHrP may also exert a systemic effect is supported
??
The Role of PTHrP in EpitheliaJ Calcium Transport
Renal Calcium Transport As evidenced by patients with HHM, PTHrP exerts both PTH-like and PTHunlike effects. According to some investigators, patients with HHM can be distinguished from those with primary hyperparathyroidism by their relative hypercalciuria, yet experimental evidence suggests that PTHrP does promote tubular reabsorption of calcium, thus contributing to the hypercalcemia in HHM (Ralston et al. 1984, Scheinman et al. 1990). This apparent paradox suggests that PTHrP, at least in HHM, may modulate renal calcium transport.
Placental Calcium Transport A clear role for PTHrP in placental calcium transport has been established through elegant studies in a thyroparathyroidectomized (TPIX) fetal sheep model. Under normal physiologic conditions, the mammalian fetus is hypercalcemic (- 13 mg/dL) relative to the mother; this fetal-maternal calcium gradient is established early in gestation. Early studies revealed that fetal parathyroidectomy resulted in a reversal of this calcium gradient. Surprisingly, infusion of PTH or maternal parathyroid extracts did not restore the gradient, whereas perfusion of the placenta with autologous fetal blood partially restored it. Infusion of fetal parathyroid extracts, partially purified PTHrP, or recombinant PTHrP results in complete restoration of the normal calcium gradient (Rodda et al. 1988). Thus, PTHrP appears to stimulate a placental calcium pump that is responsible for maintaining a relative fetal hypercalcemia during pregnancy. Using a similar in situ sheep placental perfusion model, which effectively removes the fetal skeleton as a source of calcium and enables placental calcium transport to be studied directly, Care et al. (1990) demonstrated that PTHrP (67-86) is most likely the portion of the peptide
01993, Elsevier Science Publishing Co., 1043-2760/93/$6.00
TEM Vol. 4, No. 6, 1993
responsible
for stimulating
tal calcium
and magnesium
both placen-
expand and accommodate
transport.
egg. Secondly,
Avian Oviduct Calcium Transport An additional
role for PTHrP as a regula-
tor of calcium transport is likely in avian oviducts. PTHrP is expressed only in the calcium-secreting portions of the chicken oviduct (the isthmus and shell gland) and may possibly be involved in the transfer
Calcium Transport
Finally, as discussed in some detail above, it has been suggested by some investigators that PTHrP regulates calcium transport from the maternal circulation into milk during lactation (Law et al. 1991). PTHrP as a Smooth Muscle Relaxant
PTHrP has been detected in a wide variety of types of smooth muscle, both of vascular and nonvascular origin. PTHrP appears to play a role in the mammalian reproductive process in part through its effects on uterine contractility. Studies of PTHrP protein and mRNA in rats localize PTHrP to the myometrium. Uterine PTHrP expression is induced by stretch and depends on uterine occupancy; in rats in which unilateral pregnancy was produced by tubal ligation, PTHrP expression was confined to the pregnant horn (Thiede et al. 1990). PTHrP levels peak during late gestation and drop off precipitously within 24 h of parturition. Thus, the synthesis and expression of PTHrP is regulated by stretch and, in turn, PTHrP has been shown to relax uterine smooth
cessful delivery. PTHrP appears oviduct, affecting vascular smooth gests that PTHrP muscle relaxant,
that is observed during the calcification
PTHrP plays a critical autocrine or paracrine role in normal cellular function. In certain tissues, such as the lactating mam-
speculation
that
the well-established,
PTHrP
might
but physiolog-
vasorelaxant
properties
has been shown to induce concentration-dependent relaxation in isolated rabbit renal arteries (Winquist et al. 1987) and hypotension in intact animals (Nickels et al. 1989). The physiologic role of PTHrP in vasomotor control remains unclear. Several theories regarding the vasorelaxant properties of PTHrP have been proposed. For example, as noted above, PTHrP in the avian oviduct may
of PTH.
PTHrP
allow for the increased blood flow to the shell gland seen during the calcification phase of egg laying. Similarly, it has been suggested that PTHrP may augment uterine blood flow at term. Finally, it is also possible that PTHrP produced both locally in vascular smooth muscle and systemically in the adrenal medulla (Ikeda et al. 1988) may act in conceit to regulate vascular smooth muscle tone.
to play a role in the avian both vascular and nonmuscle. Evidence sugfunctions as a smooth allowing the oviduct to
TEM Vol.4,No. 6.1993
evolutionary
ically perplexing,
orchestrate uterine relaxation with contractions, thereby helping to establish the organized rhythmicity necessary for a suc-
tissues. Both its wide-
the increased blood flow to the shell gland
share
expression should increase at term when uterine contractions are most frequent. One could propose that PTHrP serves to
in a multitude of normal as
spread tissue distribution
support
muscle. If PTHrP does indeed have a relaxant effect on the pregnant uterus, it seems counterintuitive that PTHrP gene
be expressed
well as pathologic
cular smooth muscle relaxant, allowing for
with mechanical stretch of the bladder wall (Yamamoto et al. 1992b). Exogenous PTHrP has also been shown to reverse carbachol-induced contractions of bladder muscle strips in isolated organ baths (Yamamoto et al. 1992b). Similar studies in rat gastric smooth muscle have shown that N-terminal PTHrP relaxes acetylcholine-induced muscle tension, and that this effect is blocked by the PTH receptor antagonist, PTH (334) (Mok et al. 1989). These data suggest that PTHrP mediates its smooth-musclerelaxing properties through the PTH/ PTHrP receptor. Finally, studies revealing that PTHrP is expressed by cultured rat aortic smooth muscle cells (Hong0 et al. 1991)
be elucidated.
??
as a vas-
phase of egg laying (Thiede et al. 1991). Studies on hollow viscera such as the bladder, uterus, and gastric fundus have shown that PTHrP may be intimately linked to the control of smooth muscle tonicity. PTHrP immunoreactivity and mRNA have been detected in human bladder (Thiede et al. 199 1); in experimental animals, PTHrP gene expression increases
of skeletal calcium to the eggshell during egg laying (Thiede et al. 1991). The precise molecular and cellular mechanisms of placental calcium transport remain to
Mammary
an incoming
it may function
Conclusion
Although first isolated from tumors causing HHM, PTHrP is now appreciated to
01993,
Elsevier Science Publishing Co.. 1043-2760/93/$6.00
suggest
that
mary gland, the physiologic function of PTHrP has begun to be explored. In other tissues, such as the CNS, where PTHrP is localized to specific neurons in the cerebral cortex, hippocampus, and cerebellar cortex, the physiologic role of PTHrP remains to be elucidated. PTHrP is involved in many facets of the reproductive process; it appears to regulate the growth, differentiation, and development of single cells as well as whole organisms. Its roles in calcium transport and as a smooth muscle relaxant are indisputable, yet a thorough understanding of its role in normal vascular homeostasis will require further investigation. From its discovery as the peptide responsible for HHM to the realization that PTHrP is expressed in a spectrum of normal tissues, elucidation of the true physiologic roles of PTHrP has proved to be a fascinating field of inquiry for molecular and clinical endocrinologists alike. References Adachi N, Yamaguchi Parathyroid possible
K, MiyakoY,
et al.: 1990.
hormone-related
autocrine
lymphocytes.
growth
Biochem
protein
is a
inhibitor
for
Biophys
Res Com-
mun 166:1088-1094. Albright
F: 1941. Case records
chusetts
General
of the Massa-
Hospital.
N Engl J Med
225:789-91. Allgrove J, Adami S, Manning JLH:
1985. Cytochemical
RM, O’Riordan
bioassay
of para-
thyroid hormone in material and cord blood. Arch Dis Child 60: Attillasoy
E, Burtis
11O-l15. WJ, Milstone
Immunohistochemical human
skin.
LM: 1991.
localization
thyroid hormone-related J Invest
protein Dermatol
of parain normal 96:277-
280. Broadus
AE, Stewart
hormone-related
AF: 1993. Parathyroid
protein: structure,
ing and physiological
actions.
process-
In Bilezikian
JP, Levine MA, Marcus R, eds. The Parathyroids,
Basic
and Clinical
Concepts.
New
York, Raven (in press). Brommage
??
and its intense
conservation
R, DeLuca
of bone mineral
HF: 1985. Regulation
loss during lactation.
Am J
Physiol248:E182-E187. Budayr
A, Halloran
B, King J, Diep D, Nis-
senson R, Strewler
G: 1989. High levels of a
185
ation factor: synthetic hHHF (l-34) inhibits proliferation and induces terminal differentiation of cultured human keratinocytes [abst 5821. J Bone Miner Res 3(Suppl l):S214.
calcemia of malignancy: evidence for a nonparathyroid humoral agent with an effect on renal tubular handling of calcium. Clin Sci 66:187-191.
Hongo T, Kupfer J, Enomoto H, et al.: 1991. Abundant expression of parathyroid hormone-related protein in primary rat aortic smooth muscle cells accompanies serum induced proliferation. J Clin Invest 88: 18411847.
Rodda C, Kubota M, Heath J, et al.: 1988. Evidence for a novel parathyroid hormonerelated protein in fetal lamb parathyroid glandsand sheepplacenta: comparisonswith a similar protein implicated in humoral hypercalcemia of malignancy. J Endocrinol 117:261-271.
parathyroid hormone-like protein in milk. Proc Nat1 Acad Sci USA 8617183-7185. Burtis WJ, Brady TG, Orloff JJ, et al.: 1990. Immunochemical characterization of circulating parathyroid hormone-related protein in patients with humoral hypercalcemia of cancer. N Engl J Med. 322:1106-l 112. Burton P, Moniz C, Knight D: 1990. Parathyroid hormone related peptide can function as an autocrine growth factor in human renal cell carcinoma. Biochem Biophys Res Commun 167:1134-l 138. Burton PBJ, Moniz C, Quirke P, et al.: 1992. PTHrP: expression in fetal and neonatal development. J Path01 167:291-296. Campos RV, Asa SL, Drucker DJ: 199 1. Immunocytochemical localization of PTH-LP in the rat fetus. Cant Res 51:6351-6357. Care AD, Abbas SK, Pickard DW: 1990. Stimulation of ovine placental transport of calcium and magnesium by mid-molecule fragments of human parathyroid hormone-related protein. Exp Physiol 75:605-608. Chan SD, Strewler GJ, King KL, Nissenson RA: 1990. Expression of a parathyroid hormonelike protein and its receptor during differentiation of embryonal carcinoma cells. Mol Endocrinol4:639-646. Daifotis AG, Weir EC, Dreyer BE, Broadus AE: 1992. Stretch-induced parathyroid hormonerelated peptide gene expression in the rat uterus. J Biol Chem 267:23,455-23,458. Ferguson J, Gorman J, Bruns D, et al.: 1992. Abundant expression of parathyroid hormone-related protein in human amnion and its association with labor. Proc Nat1 Acad Sci USA 89:8384-8388. Grill V, Hillary J, Ho PM, et al.: 1992. Parathyroid hormone-related protein: a possible endocrine function in lactation. Clin Endocrinol (Oxf) 37:405-410.
Insogna KL, Stewart AF, Morris CA, Hough LM, Milstone LM, Centrella M: 1989. Native and a synthetic analogue of the malignancyassociated parathyroid hormone-like protein have in vitro transforming growth factorlike properties. J Clin Invest 83:1057-1060. Juppner H, Abou-Samra AB, Freeman M, et al.: 1991. A G-protein linked receptor for parathyroid hormone and PTH-related protein. Science 254:102&1026. Kaiser SM, Laneuville P, Bemier SM, Rhim JS, Kremer R, Goltzman D: 1992. Enhanced growth of a human keratinocyte cell line induced by antisense RNA for parathyroid hormone-related protein. J Biol Chem 267:13,623-13,628. Karaplis AC, Tybulewicz Y, Mulligan RC, Kronenberg HM: 1992. Disruption of parathyroid hormone-related peptide gene leads to a multitude of skeletal abnormalities and perinatal mortality [abst 11. J Bone Miner Res 7(Suppl I):S93. Khosla S, Johansen KL, Or-y SJ, O’Brien PC, Kao PC: 1990. Parathyroid hormone-related peptide in lactation and in umbilical cord blood. Mayo Clin Proc 651408-1414.
Halloran BP, Nissenson RA: 1992. Parathyroid Hormone-Related Protein: Normal Physiology and Its Role in Cancer. Boca Raton, FL, CRC.
Law FM, MoatePJ, LeaverDD, etal.: 1991. Parathyroid hormone-related protein in milk and its correlation with bovine milk calcium. J Endocrinol 128:21-26.
Heath DA, Senior P, Beck F: 1990. Parathyroid hormone-related protein expression in the rat fetus [abst 1961. J Bone Miner Res 5(Suppl2):S122. Hillman LS, Chow W, Salmons SS, Weaver E, Erickson M, Hansen J: 1988. Vitamin D metabolism, mineral homeostasis and bone mineralization in term infants fed human milk, cow milk-based formula or soy based formula. J Pediatr 112:864-874. Hillman LS, Forte LR, Thorne PK, Johnson LS, Allen SH: 1990. Elevated PTH-related peptide in cord blood of term and premature neonates [abst 4691. J Bone Miner Res 5(Suppl 2):S191. Holick MF, Nussbaum S, Persons KS: 1988. PTH-like humoral hypercalcemic factor of malignancy may be an epidermal differenti-
186
Ikeda K, Weir EC, Mangin M, et al.: 1988. Expression of an mRNA encoding a parathyroid hormone-like peptide in normal human and animal tissues with abnormal expression in human parathyroid adenomas. Mol Endocrinol2:1230-1236.
Mok L, Eberechukwu A, Martin TJ, Thompson JC, Cooper CW: 1989. Parathyroid hormone-related protein relaxes smooth muscle and shows cross-desensitization with PTH. J Bone Miner Res 4:433-439. Moniz C, Burton P, Malik A, et al.: 1990. Parathyroid hormone-related peptide in normal human fetal development. J Mol Endocrinol 5:259-266. Nickels GA, Nana AD, Nickels MA, DiPette DJ, Asimakis GK: 1989. Hypotension and cardiac stimulation due to the parathyroid hormone-related protein, humoral hypercalcemia of malignancy factor. Endocrinology 125:834-841. Ralston SM, Fogelman I, Gardner MD, Dryburgh FJ, Cowan RA, Boyle IT: 1984. Hyper-
01993,
Elsevier Science Publishing Co., 1043-2760/93/$6.00
Scheinman SJ, Mitnick MA, Stewart AF: 1990. Direct effects of synthetic parathyroid hormone-like peptides on renal calcium transport. J Bone Miner Res 5~653-658. Schermer DT, Chan SD, Bruce RA, Wood WI, Strewler GJ: parathyroid hormone-related expression during embryologic J Bone Miner Res 6:149-155.
R, Nissenson 1991. Chicken protein and its development.
Stewart AF, Broadus AE: 1990. Humoral hypercalcemia of malignancy. In Mazzafeti E, ed. Advances in Endocrinology. Chicago, Yearbook Medical, pp l-2 1. Stewart AF, Insogna KL, Broadus AE: 1993. Malignancy- associated hypercalcemia. In De Groot L, ed. Endocrinology, 3rd ed. Philadelphia, WB Saunders (in press). Strewler GJ, Nissenson RA: 1990. Hypercalcemia in malignancy. West J Med 153:635640. Thiede M: 1989. The mRNA encoding a parathyroid hormone-like peptide is produced in mammary tissue in response to elevations in serum prolactin. Mol Endocrinol3: 14431447. Thiede M, Rodan G: 1988. Expression of a calcium-mobilizing parathyroid hormone-like peptidein lactating mammary tissue. Science 242~278-280. Thiede M, Daifotis A, Weir E, et al.: 1990. Intrauterine occupancy controls expression of the parathyroid hormone-related peptide gene in preterm rat myometrium. Proc Nat1 Acad Sci USA 87~6969-6973. Thiede MA, Harm SC, McKee RL, Grasser WA, Duong LT, Leach RM: 1991. Expression of the parathyroid hormone-related protein gene in the avian oviduct: potential role as a local modulator of vascular smooth muscle tension and shell gland motility during the egg-laying cycle. Endocrinology 129: 19581966. Van de Stolpe A, Karperien M, Clemens WG, et al.: 1993. Parathyroid hormone-related pep tide as an endogenous inducer of parietal endoderm differentiation. J Cell Biol 120:235243. Winquist RJ, Baskin EP, Vlasuk GP: 1987. Synthetic tumor- derived human hypercalcemic factor exhibits parathyroid hormone-like vasorelaxation in renal arteries. Biochem Biophys Res Commun 149:227-232.
TEM Vol. 4, No. 6, 1993
Yamamoto M, Fisher JE, Thiede MA, Caulfield MP, Rosenblatt M, Duong LT: 1992a. Concentrations of parathyroid hormone-related protein in rat milk change with duration of lactation and interval from previous suckling, but not with milk calcium. Endocrinology 130:741-747.
Yamamoto M, Harm SC, Grasser WA, Thiede MA: 1992b. Parathyroid hormone related pmtein in rat urinary bladder: a smooth muscle relaxant produced locally in response to mechanical stretch. Proc Nat1Acad Sci USA 895326-5330. TEM
Does Inhibin Have an Endocrine Function During the Menstrual Cycle? Hamish M. Fraser and Stephen F. Lunn
Inhibin (a-p heterodimer) has been considered to be the principal nonsteroidal ovarian regulator ofpituitary FSH secretion. The bfi heterodimer, activin, produced by the ovary and other tissues, appears to act locally, with actions opposite to those of inhibin. Since immunoreactive inhibin is highest during the luteal phase of the menstrual cycle when FSH is lowest, a negative feedback role in controlling FSH release at this time has been suggested. Attempts to establish this by using immunoneutralization techniques have failed to reveal such a role. We must enhance our understanding of the gonadotropic control of inhibimactivin gene expression within the various compartments of the primate ovary, the role of their binding proteins, and the nature of the secretory products before we can resolve the question of whether inhibin has an endocrine function during the menstrual cycle and how cyclic reinitiation of follicular development is controlled. (Trends Endocrinol Metab 1993;4: 187-l 94)
level. Ovarian glycoproteins,
control of FSH secretion. Of these, inhibin has been considered
ally of a single follicle, occurs mately every 28 days. Follicular
nies luteal demise, that are responsible for the cyclic reinitiation of antral follicle de-
approxidevelop-
velopment. This in turn determines the length of the menstrual cycle. It is of in-
ment is controlled by the pituitary gonadotropic hormones FSH and LH. Their se-
terest to elucidate the mechanisms by which pituitary FSH secretion is con-
cretion is driven by GnRH pulses from the hypothalamus, modulated
and it is synchronized
by a complex
back mechanisms
trolled, especially at the time of the lutealfollicular transition, because this should provide important information with re-
and
series of feed-
(Figure
1). During the
spect to the factors that trol of FSH LH in that,
luteal phase, follicles do not mature beyond
the early
antral
stage
plasma FSH concentrations generally
considered
because
are low. It is
that it is these low
levels of FSH during the luteal phase, followed by the rise in FSH that accompaHamish M. Fraser and Stephen F. Lunn are at the MRC Reproductive Biology Unit, Centre for Reproductive Biology, Edinburgh EH3 9EW, Scotland.
TEM Vol. 4,No.6,1993
role of pituitary and ovarian control folliculogenesis. Consecretion differs from that of with respect to the former, a
negative feedback component acting directly at the level of the pituitary is thought to be of major importance (Gharib et al. 1990). Ovarian estradiol is known to be the major steroid involved, although its actions are complicated by an additional effect at the hypothalamic
01993,
Elsevier Science Publishing Co., 1043-2760/93/$6.00
to be the most im-
portant. Decades of research and speculation culminated in the mid-l 980s with the characterization of inhibin as a disulfide-linked heterodimeric glycoprotein, consisting of an a-chain of 18 kD and one of two highly homologous B-chains, PA and &, of 14 kD. All three subunits are encoded by separate genes. Dimerization of the b-chains results in the formation of the activins, which exert effects opposite to those of inhibin on both pituitary FSH secretion and ovarian function (Vale et al. 1988). Classically, the biologic action of the mature inhibin molecule has been considered to be that of an endocrine factor responsible for suppressing pituitary FSH secretion, and this review examines the evidence for this role during the primate menstrual cycle. Despite extensive research on the physiology of activin and inhibin from studies in nonprimates [see A&land et al. (1992) Findlay et al. (1991), and Woodruff and Mayo (1990) for reviews], caution must be exercised when extrapolating from nonprimate models to the human, since there appear to be fundamental differences in the physiology of inhibin between primates and other species. The physiologic role of inhibin in primates has been investigated by several approaches that are discussed below.
??
During the menstrual cycle, ovulation, usu-
which prob-
ably do not cross the blood-brain barrier, are thought to provide another link in the
Secretion of Inhibin During the Menstrual Cycle
The potential endocrine role of inhibin has been examined by monitoring changes in concentrations of circulating immunoreactive inhibin throughout the menstrual cycle. Originally, inhibin was thought to be solely a product of the developing follicle, and indeed it is present in high concentrations in follicular fluid. However, the first effective RIA, based upon an antibody to purified bovine inhibin and utilizing bovine inhibin as tracer, demonstrated that the highest concentrations of immunoreactive inhibin in plasma occurred after ovulation in women (McLachlan et al. 1990, Lenton et al. 1991), Old World primates (Fraser et al. 1989, Basseti et al. 1990), and New World primates (Smith et al. 1990, Knight et al. 1992). Detailed analysis shows that in women in-
187