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Biomedical subjects

D L Stocum

Publications and source records attributed to D L Stocum.

At least 19 recordsLinked to original sources
PubMed2001

Leaping lopsided: a review of the current hypotheses regarding etiologies of limb malformations in frogs.

Recent progress in the investigation of limb malformations in free-living frogs has underlined the wide range in the types of limb malformations and the apparent spatiotemporal clustering of their occurrence. Here, we review the current understanding of normal and abnormal vertebrate limb development and regeneration and discuss some of the molecular events that may bring about limb malformation. Consideration of the differences between limb development and regeneration in amphibians has led us to the hypothesis that some of the observed limb malformations come about through misdirected regeneration. We report the results of a pilot study that supports this hypothesis. In this study, the distal aspect of the right hindlimb buds of X. laevis tadpoles was amputated at the pre-foot paddle stage. The tadpoles were raised in water from a pond in Minnesota at which 7% of surveyed newly metamorphosed feral frogs had malformations. Six percent (6 of 100) of the right limbs of the tadpoles raised in pond water developed abnormally. One truncated right limb was the only malformation in the control group, which was raised in dechlorinated municipal water. All unamputated limbs developed normally in both groups. Three major factors under consideration for effecting the limb malformations are discussed. These factors include environmental chemicals (primarily agrichemicals), encysted larvae (metacercariae) of trematode parasites, and increased levels of ultraviolet light. Emphasis is placed on the necessary intersection of environmental stressors and developmental events to bring about the specific malformations that are observed in free-living frog populations.

Animals↗
PubMed1999

Limb regeneration: re-entering the cell cycle.

Understanding the cellular plasticity that enables urodeles to regenerate many tissues is important for determining why mammals repair those same tissues with scar. The answer may lie partly in a recently discovered differential responsiveness of urodele cells to factors present in serum at the wound site.

Animals↗
PubMed1996

A conceptual framework for analyzing axial patterning in regenerating urodele limbs.

This review describes what we have learned about mechanisms of patterning in regenerating urodele limbs. Experimental evidence for three concepts is presented. First, the regeneration blastema is a self-organizing system based on positional memories inherited from parent limb cells. Second, the autonomous patterning mechanism involves local cell interactions that determine patterns of Hox gene activation. The effector molecules for positional identity reside in or on the cell surface, and can be altered by retinoic acid. Third, proximodistal patterning of the blastema is linked to blastemal growth, which in turn is dependent on a signal from the apical epidermal cap and on the non-uniformity of positional identity in the transverse axes. Lastly, the question of the degrees of similarity between the mechanisms of urodele limb regeneration, urodele limb embryogenesis, and the embryogenesis of other tetrapod limbs is discussed.

Animals↗
PubMed1993

Test of a model for the effects of retinoic acid on urodele limb regeneration.

Previous studies have shown that in axolotls (Ambystoma mexicanum), retinoic acid (RA) treatment evokes pattern completion in limb regenerates derived from anterior and dorsal half zeugopodia (lower arms and legs), but causes regenerative failure in posterior and ventral half zeugopodia. Pattern completion in anterior and dorsal half limbs may be explained by postulating that intercalary regeneration occurs in the antero-posterior (AP) and dorsoventral (DV) axes between blastema cells that are posteriorized (anterior half limb) or ventralized (dorsal half limb) by RA, and circumferential anterodorsal cells that remain unaffected by RA and thus maintain their original positional identities. The contrasting regenerative failure of RA-treated posterior and ventral half zeugopodia may likewise be explained by postulating that all the blastema cells in the posterior half are posteriorized, and all the cells in the ventral half are ventralized by RA, thus eliminating differentials in transverse positional identity essential for blastema formation and outgrowth. To test these postulates we grafted blastemas derived from limbs halved in the AP and DV axes of control and RA-treated animals to untreated whole limb stumps and analyzed the patterns of supernumerary (SN) regeneration. The site or location of SN formation will demonstrate (1) whether RA has posteriorized and ventralized the positional identity of the blastema cells and (2) if blastema cells in the periphery of the anterodorsal quadrant of the limb are resistant to these RA-induced changes in positional identity.(ABSTRACT TRUNCATED AT 250 WORDS)

Ambystoma↗
PubMed1992

Retinoic acid-induced change in anteroposterior positional identity in regenerating axolotl limbs is dose-dependent.

Retinoic acid (RA) induces pattern duplication in the proximodistal (PD) axis of axolotl limb regenerates. The effect is dose-dependent, with the maximum extent of duplication being evoked at a dose of 150 micrograms RA/g body weight. The same dose of RA induces maximum pattern completion in the anteroposterior (AP) axis of regenerating anterior half or double anterior half limbs. RA inhibits the regeneration of posterior half or double posterior half limbs (Kim, W.S., and Stocum, D.L. [1986] Dev Biol 114:170-179). The effects of lower doses of RA on the AP axis of limb regenerates have not been tested and thus it is not known whether the effects of RA on positional identity in this axis are dose-dependent or are all-or-none. To answer this question, we examined the effects of a range of doses of RA on AP positional identity in regenerating double anterior and double posterior axolotl forelimbs and hindlimbs constructed by exchanging the anterior and posterior halves of right and left zeugopodia as either autografts or homografts. Ten days after the exchange, the double half zeugopodia were amputated through their distal ends. At 4 days postamputation, the animals were injected intraperitoneally with 20, 50, 75, or 100 micrograms RA/g body weight and the constructs allowed to regenerate for at least 6 weeks. Control double anterior forelimbs and hindlimbs formed symmetrical regenerates with an average of two and 1.8 anterior digits, respectively. RA treatment induced the blastema of double anterior zeugopodia to regenerate missing posterior structures in mirror-image patterns and to duplicate zeugopodial and stylopodial elements in the PD axis.(ABSTRACT TRUNCATED AT 250 WORDS)

Ambystoma↗
PubMed1990

The effect of retinoic acid on positional memory in the dorsoventral axis of regenerating axolotl limbs.

We investigated the effect of retinoic acid (RA) on pattern regulation in the dorsoventral (DV) axis of regenerating axolotl limbs. Half and double half dorsal and ventral zeugopodia (lower arms or legs) were amputated through their distal ends, and 4 days later the animals were injected intraperitoneally with 50 (large animals) or 100 (small animals) micrograms RA/g body wt. Half and double half dorsal and ventral zeugopodia of uninjected axolotls, and sham-operated zeugopodia of untreated and RA-treated limbs served as controls. Skeletal patterns and the DV muscle patterns of control and experimental regenerates were then analyzed. Sham-operated zeugopodia of uninjected animals regenerated normally. Sham-operated, RA-treated zeugopodia regenerated normally with proximodistal duplications. Sixty percent of uninjected control dorsal half zeugopodia, 80% of control ventral half zeugopodia, and 100% of control double dorsal and double ventral zeugopodia regenerated distally, but the regenerates did not reconstitute the muscle pattern of the missing half. Thirty-eight percent of RA-treated ventral half zeugopodia and 78% of RA-treated double ventral zeugopodia failed to regenerate distally. Of those cases that did regenerate distally, none regenerated the muscle pattern of the missing half. By contrast, 100% of RA-treated dorsal half zeugopodia regenerated distally and all completed the normal DV muscle pattern. Forty-one percent of RA-treated double dorsal zeugopodia failed to regenerate, but of the remainder that did regenerate, 50% completed the normal DV muscle pattern. These represented eight cases, six of which regenerated single limbs, and two of which regenerated twin limbs, each with a normal DV muscle pattern. We interpret these data to mean that RA ventralizes the positional memory of blastema cells in the DV axis.

Ambystoma↗
PubMed1988

Cellular retinoic acid binding protein: detection and quantitation in regenerating axolotl limbs.

The concentrations of apo (unoccupied), holo (occupied), and total cellular retinoic acid binding protein (CRABP) were measured at various stages of axolotl limb regeneration. The ratio of apo-CRABP to holo-CRABP declined with advancing regenerate stage until the CRABP was all in the holo form. The increase in holo-CRABP is correlated with a stage-dependent shift in the effect of exogenous retinoic acid on regenerate pattern, from pattern duplication to inhibition of regeneration. The data suggest, though they do not prove, that these different morphological effects could be due to a shift from a CRABP-dependent to a CRABP-independent mechanism of exogenous retinoic acid (RA) action that is related to stage-specific variations in endogenous RA levels.

Ambystoma↗
PubMed1988

Retinoic acid coordinately proximalizes regenerate pattern and blastema differential affinity in axolotl limbs.

An assay that detects position-related differences in affinity of axolotl regeneration blastema cells in vivo was used to test whether retinoic acid, which proximalizes regenerate pattern, simultaneously proximalizes blastema cell affinity. The assay involved autografting or homografting late bud forelimb blastomas derived from the wrist, elbow or midupper arm levels to the dorsal surface of the blastema-stump junction of an ipsilateral, medium-bud-stage hindlimb regenerating from the midthigh level. The grafted blastemas consistently displaced to their corresponding levels on the proximodistal axis of the host regenerate, indicating the existence of level-specific differences in blastema cell affinity. Retinoic acid proximalized the pattern of donor forelimb regenerates to the level of the girdle and abolished their displacement behaviour on untreated host hindlimbs. Conversely, untreated forelimb donor blastemas displaced distally to their corresponding levels on host ankle regenerates, that had been proximalized to the level of the girdle by retinoic acid. These results indicate that positional memory in regenerating limbs is directly related to blastema cell affinity, and that very similar or identical sets of level-specific affinity properties are shared by forelimb and hindlimb cells.

Ambystoma↗
PubMed1988

Retinoic acid proximalizes level-specific properties responsible for intercalary regeneration in axolotl limbs.

The objective of this study was to determine whether retinoic acid (RA) coordinately proximalizes positional memory and the cellular recognition system that detects pattern discontinuity in regenerating amphibian limbs. The strategy was to test the capacity of RA-treated blastemas to evoke intercalary regeneration when grafted to an amputation level proximal to their level of origin. Control wrist and ankle, or elbow and knee blastemas treated with the retinoid solvent, dimethylsulphoxide, evoked intercalary regeneration as effectively as untreated blastemas, when grafted to the midstylopodial amputation surface of host limbs. RA-treated wrist and ankle or elbow and knee blastemas were proximalized and formed complete limbs that were at an angle to, or continuous with, the midstylopodium of the host limb. No intercalary regeneration, from either graft or host, was observed in these cases. The results indicate that the cellular mechanism that recognizes disparities between non-neighbouring cells and initiates intercalary regeneration is coordinately proximalized with positional memory. Thus the recognition mechanism and positional memory are directly related. Intercalary regeneration and corrective displacement (affinophoresis), both of which restore a pattern of normal cell neighbours by different means in regenerating axolotl limbs, appear to use the same mechanism to recognize pattern discontinuity.

Ambystoma↗
PubMed1987

Use of retinoids to analyze the cellular basis of positional memory in regenerating amphibian limbs.

Cells of the amphibian limb regeneration blastema inherit memories of their level of origin (positional memory) along the limb axes. These memories serve as boundaries of what is to be regenerated, thus preventing regeneration of any but the missing structures. Because of its importance in determining the boundaries of regenerate pattern, it is essential to understand the cellular and molecular basis of positional memory. One approach to this problem is to look for position-related differences in a cell or molecular property along a limb axis and then show, using an agent that modifies regenerate pattern, that the cell or molecular property and the pattern are coordinately modified. We have done this using retinoic acid (RA) as a pattern-modifying agent and an in vivo assay that detects position-related differences in a cell recognition-affinity property along the proximodistal (PD) axis of the regenerating axolotl limb. RA proximalizes positional memory in the PD axis, posteriorizes it in the anteroposterior axis, and ventralizes it in the dorsoventral axis. The level-specific PD cell recognition-affinity property is proximalized by RA, indicating that this property and positional memory are causally related. The effects of RA on positional memory may be mediated through a cellular RA-binding protein (CRABP), since the concentration of unbound (apo) CRABP molecules is highest during early stages of regeneration when the proximalizing effects of RA are greatest.

Amphibians↗
PubMed1986

Retinoic acid modifies positional memory in the anteroposterior axis of regenerating axolotl limbs.

The effects of retinoic acid (RA) on anteroposterior (AP) positional memory of regenerating axolotl limbs were tested after removing the anterior or posterior half from the zeugopodium (lower arm or leg). RA (150 micrograms/g body wt) was injected into groups of animals bearing the following types of limbs: (1) anterior and posterior half zeugopodia grafted to the eyesocket and amputated distally 7 days later; (2) unamputated anterior and posterior half zeugopodia in situ; (3) double anterior and double posterior half zeugopodia amputated distally 7 days after their construction; (4) sham-operated zeugopodia amputated distally 7 days after operation. Controls consisted of these four groups injected with the retinoid solvent, dimethyl sulfoxide, or not injected. Control half zeugopodia grafted to the eyesocket regenerated no more than one or two digits. Control unamputated half zeugopodia in situ underwent partial or complete regeneration of the missing half from the proximal and midline wound surfaces exposed during construction of the half zeugopodia. Control double anterior and posterior zeugopodia both regenerated symmetrical, hypomorphic regenerates with 1-3 digits in the double anteriors and 1-6 digits in the double posteriors. Sham-operated controls regenerated normally. Regenerating anterior and posterior halves responded differently to RA. RA-treated anterior half zeugopodia in the eyesocket, and anterior half stumps adjacent to the unamputated posterior half zeugopodia in situ both produced regenerates that duplicated stump structures in the proximodistal axis and formed a complete and normal AP pattern. RA-treated double anterior zeugopodia regenerated proximodistal-duplicated pairs of mirror-imaged limbs, each with a complete and normal AP pattern. In contrast, half posterior zeugopodia in the eyesocket, the posterior half stumps of unamputated half anterior zeugopodia in situ, and double posterior zeugopodia all failed to regenerate. These results suggest that RA modifies positional memory in only one direction in the AP axis, posterior.

Ambystoma↗